Purine diones as wnt pathway modulators

ABSTRACT

The invention relates to the use of compounds of general structure (I) in modulation of the Wnt pathway [Formula should be inserted here] wherein R 1 , R 2 , R 3 , R 4  and R 5  are each, independently, H or an alkyl group; D is selected from the group consisting of H, halogen, alkyl, cycloalkyl, aryl, and dialkylamino, each (other than H and halogen) being optionally substituted; Ar is an aryl or heteroaryl group, optionally substituted; Cy is an aryl, heteroaryl or a saturated ring containing at least one heteroatom, each being optionally substituted; and n is an integer from 1 to 3.

FIELD

The invention relates to Wnt pathway modulators, processes for makingthem and methods for using them.

BACKGROUND

The present application claims priority from application GB1309333.1,filed on 23 May 2013, the entire contents of which are incorporatedherein by cross-reference.

Wnt proteins are secreted glycoproteins acting as growth factors thatregulate various cellular functions, including proliferation,differentiation, death, migration, and polarity, by activating multipleintracellular signalling cascades, including the β-catenin-dependent and-independent pathways. There are 19 Wnt members that have been found inhumans and mice, and they exhibit unique expression patterns anddistinct functions during development. In humans and mice, the 10members of the Frizzled (Fz) family comprise a series of seven-passtransmembrane receptors that have been identified as Wnt receptors. Inaddition to Fz proteins, single-pass transmembrane proteins, such aslow-density lipoprotein receptor-related protein 5 (LRP5), LRP6,receptor tyrosine kinase (RTK)-like orphan receptor 1 (Ror1), Ror2, andreceptor-like tyrosine kinase (Ryk), have been shown to function asco-receptors for Wnt signalling. Therefore, it has been assumedtraditionally that the binding of different Wnts to their specificreceptors selectively triggers different outcomes via distinctintracellular pathways.

In the absence of Wnt signalling, β-catenin is bound and phosphorylatedby a “destruction complex” containing the adenomatous polyposis coli(APC) and Axin proteins, as well as glycogen synthase kinase 3 (GSK3)and casein kinase I (CKI). Phosphorylated β-catenin is bound by the Fbox protein Slimb/β-TrCP and polyubiquitinated, leading to proteosomaldegradation. In addition, the complex acts to prevent nuclearlocalization of β-catenin. Upon Wnt binding to Frizzled (Fz) andlow-density lipoprotein-related proteins 5 and 6 (LRP5/6), GSK3, Axin,and other destruction complex components are recruited to the receptorcomplex. The function of the destruction complex is inhibited, andunphosphorylated β-catenin accumulates in the cytoplasm and eventuallytranslocates to the nucleus. There, it associates with TCF proteins,converting TCF from a repressor into an activator of Wnt-responsive genetranscription.

Deregulation of components of Wnt/β-catenin signalling is implicated ina wide spectrum of diseases including degenerative diseases, metabolicdiseases, and a number of cancers such as cervical, colon, breast,bladder, head and neck, gastric, lung, ovarian, prostate, thyroid,non-small-cell lung, as well as chronic lymphocytic leukemia,mesothelioma, melanoma, pancreatic adenocarcinoma, basal cell carcinoma,osteosarcoma; hepatocellular carcinoma, Wilm's tumor andmedulloblastoma. Wnt signalling plays a role both during development,and within stem cell niches in adults. This is best established in skin,hematopoietic stem cells, mammary gland and in intestinal proliferation.For example, high level expression of DKK1, an inhibitor of Wntsignalling, blocks normal stem cell proliferation in mouse intestines,suggesting there is an essential role for Wnt signalling in maintenanceof stem cells in the digestive tract. The role of Wnt in self renewaland expansion of stem cells has also been demonstrated for embryonic andneural stem cells, suggesting that Wnt signalling may be a generalrequirement of stem cell maintenance.

Inhibition of Wnt signalling, e.g., by overexpression of axin or anextracellular Wnt-binding protein, sFRP, reduces hematopoietic stem cell(HSC) growth in vitro and the ability to reconstitute HSCs in vivo.Notably, while overexpression of activated β-catenin can expand HSCpopulations in culture for extended periods, two groups have reportedthat β-catenin is not required for HSC survival and serialtransplantation, supporting the proposal that there is more to Wntsignalling than stabilization of β-catenin in stem cell survival.Diverse Wnts can regulate stem cell proliferation: Wnts 1, 5a, and 10bare able to stimulate expansion of HSC populations and Wnt5a actssynergistically with stem cell factor (SCF) to expand and promoteself-renewal of HSCs. The demonstration of a role for Wnt5a in HSCself-renewal and its ability to synergize with stem cell factor isparticularly interesting because Wnt5a often acts in a β-cateninindependent manner. While Wnt signalling is critical for stem cellmaintenance, it may therefore be via signalling pathways distinct fromor in parallel to the β-catenin pathway.

Wnt/β-catenin signalling pathway is essential to embryonic developmentin general and organ morphogenesis, so it is not surprising thatdysregulation of this pathway in adult has been linked to fibroblastbiology and fibrosis. It has been demonstrated that Wnt/β-cateninsignalling play a role in severe fibrotic diseases, such as pulmonaryfibrosis, liver fibrosis, skin fibrosis and renal fibrosis.

Dysregulation of Wnt/β-catenin signalling contributes also to thedevelopment of diabetic retinopathy by inducing retinal inflammation,vascular leakage, and neovascularization. The binding of Wnt proteins toplasma membrane receptors on mesenchymal cells induces thedifferentiation of these cells into the osteoblast lineage and therebysupports bone formation. Wnts are also key signalling-proteins in jointremodeling processes. Active Wnt signalling contributes to osteophyteformation and might have an essential role in the anabolic pattern ofjoint remodeling that is observed in ankylosing spondylitis andosteoarthritis. In contrast, blockade of Wnt signalling facilitates boneerosion and contributes to catabolic joint remodeling, a process that isobserved in rheumatoid arthritis.

There is therefore a need for compounds that modulate or inhibit Wntactivity so as to treat diseases associated with Wnt activity.

SUMMARY OF INVENTION

In a first aspect of the invention there is provided a compound ofstructure (I) for use in, or when used in, modulating Wnt activity,

wherein:

R¹, R², R³, R⁴ and R⁵ are each, independently, H or an alkyl group;

D is selected from the group consisting of H, halogen, alkyl,cycloalkyl, aryl, and dialkylamino, each (other than H and halogen)being optionally substituted;

Ar is an aryl or heteroaryl group, each being optionally substituted;

Cy is an aryl, heteroaryl or a saturated ring containing at least oneheteroatom, each being optionally substituted; and

n is an integer from 1 to 3.

In some embodiments, if n=1 and one of R³ and R⁴ is methyl and the otheris H, the stereochemistry of the compound is as shown in partialstructure (II)

In some particular embodiments, n=1 and one of R³ and R⁴ is methyl andthe other is H and the stereochemistry of the compound is as shown inpartial structure (II).

Any one or more of the following groups a) to e), and/or any one or moreindividual compounds within any one or more of said groups, optionallyall thereof, may be excluded from the scope of the first aspect.Notwithstanding any such exclusions, the compounds referred tohereinafter as compound 5 and compound 86 may be explicitly includedwithin the scope of the first aspect.

a) Any one or more of the compounds described in WO2010/036821 A1;b) Any one or more compounds of the formula shown below,

wherein, R¹ and R² are each independently C₁-C₆ alkyl, C₂-C₆ alkenyl, orC₂-C₆ alkynyl, each of which is optionally substituted with 1-4 R⁵; L isNR⁶SO₂, SO₂NR⁶, OC(O)NR⁶, NR⁶C(O)O, NR⁶C(O)NR⁶, NR⁶C(O), C(O)NR⁶, O,C(O), S, S(O), S(O)₂, NR₆, or CH₂, each of R^(3a) and R^(3b) isindependently cyclyl, heterocyclyl, aryl, heteroaryl, each of which isoptionally substituted with 1-4 R⁷; each R⁵ is independently halo,hydroxyl, alkoxy, amino, alkylamino, dialkylamino, cyano, nitro, amido(e.g., where the nitrogen of the amide is substituted by an alkyl, orwhere the nitrogen of the amide together with two carbons to which it isattached, forms a ring), alkylamido, dialkylamido, thioyl, sulfonyl,cyclyl, heterocyclyl, aryl, or heteroaryl; each R⁶ is independently H,C₁-C₆ alkyl, C₁-C₆ alkenyl, hydroxyC₁-C₆ alkyl, alkoxyC₁-C₆ alkyl,cyanoalkyl, haloalkyl, arylalkyl, S(O)alkyl, acyl, amino, amidyl, orS(O)₂H, aryl, alkoxyaryl; each R⁷ is independently C₁-C₆ alkyl, C₂-C₆alkenyl, or C₂-C₆ alkynyl, halo, hydroxyl, alkoxy, oxo, aryl,heteroaryl, cyclyl, heterocyclyl, arylalkyl, heteroarylalkyl,cyclylalkyl, heterocyclylalkyl, aryloxy, arylalkoxy, amino, akylamino,dialkylamino, thioyl, alkylthioyl, sulfonyl, sulfonamidyl, amido (e.g.,where the nitrogen of the amide is substituted by an alkyl, or where thenitrogen of the amide together with two carbons to which it is attached,forms a ring), hydroxyl alkoxyl, alkoxy —C(O)OH, —C(O)Oalkyl, urea,sulfonylurea, acyl, nitro, cyano, each of which is optionallysubstituted with 1-3 R⁸; each R⁸ is independently C₁-C₆ alkyl, C₂-C₆alkenyl, or C₂-C₆ alkynyl, aryl, heteroaryl, cyclyl, halo, hydroxyl,alkoxy, oxo, aryloxy, amino, akylamino, dialkylamino, C(O)OH,—C(O)Oalkyl, thioyl, sulfonyl, sulfonamidyl, amido (e.g., where thenitrogen of the amide is substituted by an alkyl, or where the nitrogenof the amide together with two carbons to which it is attached, forms aring), urea, sulfonylurea, acyl, nitro, cyano, cyclyl, heterocyclyl,aryl, or heteroaryl; R⁹ is H, C₁-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆alkynyl, halo, C₁-C₆ haloalkyl, hydroxyl, alkoxy, aryloxy, arylalkoxy,amino, akylamino, dialkylamino, thioyl, alkylthioyl, sulfonyl,sulfonamidyl, amido, urea, sulfonylurea, acyl, nitro, cyano, each ofwhich is optionally substituted with 1-3 R⁸; m is 1, 2, 3, 4, 5, or 6.c) Any one or more compounds of structure I wherein R¹ and R² are bothmethyl, D, R³, R⁴ and R⁵ are all H, n is 1 and Ar-Cy is4-phenylthiazole-2-yl, 4-(4-bromophenyl)thiazole-2-yl, 2- or4-(N-morpholiniyl)phenyl, 4-(N-piperidinyl)phenyl,1-phenyl-3-methylpyrazole-5-yl, 4-(3,4-dimethylphenyl)thiazole-2-yl,4-(4-ethylphenyl)thiazole-2-yl, 4-(2,4,6-trimethylphenyl)thiazole-2-yl,4-(4-n-propylphenyl)thiazole-2-yl,4-(4-chorophenyl)-5-methylthiazole-2-yl, 4-phenylphenyl,2-(N-pyrrolidinyl)phenyl, 4-(N-azacycloheptyl)phenyl,4-(4-bromophenyl)-5-trifluoromethylpyrimidine-2-yl,4-(2,4-dimethylphenyl)thiazole-2-yl,4-(3,4-dimethoxylphenyl)thiazole-2-yl, 4-(2-fluorophenyl)thiazole-2-yl,4-(3,4-difluorophenyl)-5-methylthiazole-2-yl,4-(4-methoxy-1-naphthyl)thiazole-2-yl,4-(3-chloro-4-methoxyphenyl)thiazole-2-yl,4-(4-ethoxyphenyl)-5-methylthiazole-2-yl,4-(2-acetamido-4-fluorophenyl)thiazole-2-yl,4-(3-fluoro-4-methoxyphenyl)thiazole-2-yl,4-(4-bromophenyl)-5-methylthiazole-2-yl,4-(4-(2-mnethoxyethyl)phenyl)thiazole-2-yl,4-(4-methyl-3-nitrophenyl)thiazole-2-yl,4-phenyl-5-(1-methyl-imidazole-2-yl)thiazole-2-yl,4-(4-(difluromethoxy)phenyl)-5-methylthiazole-2-yl,4-(2,5-diethoxyphenyl)thiazole-2-yl,4-(4-fluoromethylphenyl)-5-methylthiazole-2-yl,4-phenyl-5-acetylthiazole-2-yl,4-(3-chloro-4-fluorophenyl)thiazole-2-yl,4-(3-fluorophenyl)thiazole-2-yl, 4-(3-methoxyphenyl)thiazole-2-yl,4-(2-ethoxyphenyl)thiazole-2-yl, 4-(3-chlorophenyl)thiazole-2-yl,4-(2-methoxyphenyl)thiazole-2-yl, 4-(2-bromophenyl)thiazole-2-yl,4-(4-diethylaminophenyl)thiazole-2-yl, 4-(2-chlorophenyl)thiazole-2-yl,4-(4-methoxyphenyl)thiazole-2-yl, 4-(3-bromophenyl)thiazole-2-yl,4-(4-isopropoxyphenyl)thiazole-2-yl,4-(2,3,4-trichlorophenyl)thiazole-2-yl,4-(3-chloro-4-ethoxyphenyl)thiazole-2-yl,4-(4-chloro-3-fluorophenyl)thiazole-2-yl,4-(3-ethoxyphenyl)thiazole-2-yl, 4-(2, 4-dimethoxyphenyl)thiazole-2-yl,4-(2, 4-dimethylphenyl)thiazole-2-yl, 4-(4-ethylphenyl)thiazole-2-yl,4-(2-methyphenyl)thiazole-2-yl, 4-(2-bromo-5-ethoxyphenyl)thiazole-2-yl,4-(5-bromo-2-chloro-4-ethoxyphenyl)thiazole-2-yl,4-(2-chloro-4-ethoxyphenyl)thiazole-2-yl, 4-(3-pyridyl)thiazole-2-yl,4-(4-(2-hydroxyethoxy)phenyl)thiazole-2-yl,4-(3-(2-hydroxyethoxy)phenyl)thiazole-2-yl,4-(2,6-dichlorophenyl)thiazole-2-yl, 4-(3-methylphenyl)thiazole-2-yl,4,5-bis(4-methylphenyl)oxazole-2-yl, 4,5-diphenyloxazole-2-yl,4-(4-pyridyl)thiazole-2-yl, 4-(2-(2-hydroxyethoxy)phenyl)thiazole-2-yl,4-(3,4-dichlorophenyl)-5-methylthiazole-2-yl,4-(2,4,6-trichlorophenyl)thiazole-2-yl,4-(4-ethoxyphenyl)-5-phenylthiazole-2-yl,4-(4-chloro-3-ethoxyphenyl)thiazole-2-yl,4-(3,4,5-trichlorophenyl)thiazole-2-yl, 5-(4-pyridyl)1,3,4-thiadiazole-2-yl,4-(3-bromo-6-(2-hydroxyethoxy)phenyl)thiazole-2-yl,3-(benzoxazole-2-yl)-2-methylphenyl,4-(3,4-dichlorophenyl)-5-ethylthiazole-2-yl,4-((3-trifluoromethyl)phenyl)thiazole-2-yl,4-(2,3-dichlorophenyl)-5-phenylthiazole-2-yl,4-(2,3-dichlorophenyl)-5-(4-methylphenyl)thiazole-2-yl,1-phenyl-3-(4-methylphenyl)pyrazole-5-yl,4,5-bis(3,4-dichlorophenyl)thiazole-2-yl, 5-phenylthiazole-2-yl,4-(4-(trifluoromethyl)phenyl)thiazole-2-yl,4-(4-(methylsulfinyl)phenyl)thiazole-2-yl,4-(4-chlorophenyl)oxazole-2-yl,4-((4-trifluoromethoxy)phenyl)thiazole-2-yl, 3-(4-,chlorophenyl)pyrazole-5-yl, 3-(4-chlorophenyl) 1,3,4-oxadiazole-5-yl,3-phenylpyrazole-5-yl, 5-(4-methylphenyl)thiazole-2-yl,4-(4-chlorophenyl)-5-ethylthiazole-2-yl, 4-(4-nitrophenyl)thiazole-2-yl,4-(4-aminophenyl)thiazole-2-yl, 4-(4-ethoxyphenyl)-5-methyloxazole-2-yl,4-(4-dim ethyl amino-3-fluorophenyl)thiazole-2-yl,3-(thiophene-2-yl)pyrazole-5-yl, 3-(furan-2-yl)pyrazole-5-yl,4-(4-chlorophenyl)-5-fluorothiazole-2-yl, 2-(pyrrole-N-yl)phenyl,2-(indole-2-yl)phenyl, 4-(2-hydroxybenzimidazole-5-yl)thiazole-2-yl,2-(4-chlorophenyl)-4-pyridyl, 3-(4-chlorophenyl)phenyl,4-(3-chloro-4-dimethylaminophenyl)thiazole-2-yl,4-(4-ethoxyphenyl)thiazole-2-yl, 5-(4-chlorophenyl)-3-pyridyl, 4-(3,4-dichlorophenyl)-5-isopropylthiazole-2=yl,4-(4-dimethylamino-2-fluorophenyl)thiazole-2-yl,4-(4-(tetrahydropyrrole-N-yl)-3-bromophenyl)thiazole-2-yl,4-(4-fluoro-3-(morpholine-N-yl)phenyl)thiazole-2-yl,5-(4-chlorophenyl)pyrazole-2-yl, 4-(2,4,5-trichlorophenyl)thiazole-2-yl,4-(4-diethylaminophenyl)thiazole-2-yl,4-(2,3-dihydrobenzofuran5-yl)thiazole-2-yl,2-(3,4-dichlorophenyl)thiazole-4-yl,3-phenyl-4-(4-ethoxyphenyl)imidazole-2-yl,4-(piperidine-N-ylphenyl)thiazole-2-yl,4-(morpholine-N-ylphenyl)thiazole-2-yl,4-(3-diethylaminophenyl)thiazole-2-yl, 2-(benzothiazole-2-yl)phenyl,4-(3-ethylpropylaminophenyl)thiazole-2-yl,4-(3-dimethylaminophenyl)thiazole-2-yl, 4-(4-chlorophenyl)pyridine-2-yl,3-(4-diethylaminophenyl)phenyl, 4-(4-ethoxyphenyl)imidazole-2-yl,4-(N-acetyl-2,3-dihydroindole-5-yl)thiazole-2-yl,4-(3,4-dichlorophenyl)pyrimidine-2-yl, 3-(thiazole-2-yl)phenyl,4-(quinoline-2-yl)thiazole-2-yl, 4-(benzothiophene-2-yl)thiazole-2-yl,4-(isoquinoline-3-yl)thiazole-2-yl,4-(4-dimethylamino-3,5-difluorophenyl)thiazole-2-yl,4-(3-(pyrrolidine-N-yl)phenyl)thiazole-2-yl,4-(4-(aminocarbonylamino)phenyl)thiazole-2-yl,4-(4-chlorophenyl)-5-methoxythiazole-2-yl,4-(4-diethylsulfonamidophenyl)thiazole-2-yl,4-(4-dimethylsulfonamidophenyl)thiazole-2-yl,4-(2,3-dihydro-1,4-benzoxazine-3-one-6-yl)thiazole-2-yl,4-(3,4-dichlorophenyl)imidazole-2-yl, 4-(4-ethoxyphenyl)oxazole-2-yl,4-(4-chlorophenyl)oxazole-2-yl, 3-(4-ethylthiazole-2-yl)phenyl,3-(5-ethylthiazole-2-yl)phenyl, 3-(4-methylthiazole-2-yl)phenyl,4-(2-chloro-4-diethylaminophenyl)thiazole-2-yl,4-(2,5-difluoro-4-diethylaminophenyl)thiazole-2-yl,4-(3-trifluoromethoxyphenyl)thiazole-2-yl,6-(4-chlorophenyl)pyridine-2-yl,3-(4-diethylamino-3-fluorophenyl)phenyl,4-(4-dimethylamino-2,3-difluorophenyl)thiazole-2-yl),4-(4-(piperidine-N-ylsulfonyl)phenyl)thiazole-2-yl,4-(N-acetylindole-5-yl)thiazole-2-yl,1-phenylmethyl-4-(3,4-dichlorophenyl)imidazole-2-yl,4-(N-ethyl-2,3-dihydroindole-5-yl)thiazole-2-yl, 3-(4-chlorophenyl)1,2,4-thiadiazole-5-yl,4-(N-acetyl-2,3-dihydroindole-5-yl)thiazole-2-yl,4-(N-ethylindole-5-yl)thiazole-2-yl, 1-(4-chlorophenyl)1,2,4-triazole-3-yl, 1-phenylpyrazole-4-yl,4-(N-acetylindole-6-yl)thiazole-2-yl, 1-(4-chlorophenyl)pyrazole-4-yl,4-(N-methyl-2,3-dihydroindole-5-yl)thiazole-2-yl,4-(2-chloro-4-dimethylamino-5-fluorophenyl)thiazole-2-yl,4-(3-(N,N-dimethylsulfonamido)thiazole-2-yl,4-(3-(N,N-diethylsulfonamido)thiazole-2-yl, 2-(4-chlorophenyl)1,3,5-triazine-6-yl, 4-(4-(N-morpholino)sulfonylphenyl)thiazole-2-yl,4-(4-(N-pyrrolidino)sulfonylphenyl)thiazole-2-yl,4-(4-chloro-3-diethylaminophenyl)thiazole-2-yl,4-(4-chloro-3-trifluoromethylphenyl)thiazole-2-yl,1-ethyl-4-(3,4-dichlorophenyl)imidazole-2-yl,1-methyl-4-(3,4-dichlorophenyl)imidazole-2-yl, 2-(1-naphthyl)-6-pyridyl,2-(1-(3,4-dichlorophenyl)-6-pyridyl,4-(4-chlorophenyl)-5-dimethylaminomethylthiazole-2-yl,4-(3-diethylaminophenyl)thiazole-2-yl,4-(3-fluoro-4-diethylaminophenyl)-5-ethylthiazole-2-yl,4-(4-chlorophenyl)-5-(2-dimethylaminoethyl)thiazole-2-yl,4-(2-fluoro-4-trifluoromethylphenyl)thiazole-2-yl,4-(3-fluoro-5-trifluoromethylphenyl)thiazole=2-yl,4-(3-fluoro-4-trifluoromethylphenyl)thiazole-2-yl,4-(4-fluoro-3-trifluoromethylphenyl)thiazole-2-yl,4-(3,4-dichlorophenyl)-5-methoxycarbonylmethylthiazole-2-yl,1,5-dimethyl-4-(3,4-dichlorophenyl)imidazole-2-yl,N-(4-chlorophenyl)pyrrole-3-yl,4-(3,4-dichlorophenyl)-5-aminocarbonylthiazole-2-yl,4-(2-ethoxynaphth-6-yl)thiazole-2-yl,4-(3,4-dichlorophenyl)-5-hydroxycarbonylmethylthiazole-2-yl,4-(4-diethylamino-2,5-difluorophenyl)-5-methylthiazole-2-yl,4-(4-(N-pyrrolidino)-3-fluorophenyl)thiazole-3-yl,4-(4-diethylamino-3-ethoxyphenyl)thiazole-3-yl,4-(4-diethylamino-2-ethoxyphenyl)thiazole-3-yl,4-(4-(N-pyrrolidino)-3-trifluoromethylphenyl)thiazole-3-yl,4-(4-diethylamino-3-trifluoromethylphenyl)thiazole-3-yl,1-n-butyl-4-(3,4-dichlorophenyl)imidazole-2-yl,1-(4-chlorophenyl)imidazole-4-yl, 5-(4-chlorophenyl)1,2,4-oxadiazole-3-yl, 4-phenyl-5-(4-ethoxyphenyl)oxazole-2-yl,2-(2-naphthyl)pyridine-6-yl, 4-(4-methoxycarbonylphenyl)thiazole-2-yl,4-(3-(methylethylamino)phenyl)-5-methylthiazole-2-yl,4-(3-diethylamino-4-ethoxyphenyl)thiazole-2-yl,4-(4-diethylaminophenyl)-5-fluorothiazole-2-yl,4-(4-(N-ethylpiperazine-N′-yl)-3-trifluoromethylphenyl)thiazole-2-yl,4-(3-ethoxycarbonylphenyl)thiazole-2-yl,4-(3,5-difluoro-4-(N-pyrrolidino)phenyl)thiazol-2-yl,4-(4-chlorophenyl)-5-trifluoromethylthiazole-2-yl,4-(3-cyano-4-(methylethylamino)phenyl)thiazole-2-yl,4-(2-ethoxynaphth-5-yl)thiazole-2-yl,5-(4-chloromethylphenyl)isothiazole-3-yl,4-(N-methylindole-5-yl)thiazole-2-yl,4-(3,4-trichlorophenyl)-5-(2-hydroxyethyl)thiazole-2-yl,2-(4-chlorophenyl)imidazole-4-yl, 1-(4-chlorophenyl)1,2,3-thiazole-4-yl, 4-(4-(N-imidazolyl)phenyl)thiazole-2-yl,4-(4-(N-tetrazolyl)phenyl)thiazole-2-yl,4-(4-(4-methoxyphenyl)methylaminophenyl)thiazole-2-yl,4-(4-(N-pyrrolidinyl)-3,5-difluorophenyl)-5-fluorothiazole-2-yl,4-(4-(N-morpholino)-3-trifluoromethylphenyl)thiazole-2-yl,4-(4-(N-piperidinyl)-3-trifluoromethylphenyl)thiazole-2-yl,4-(4-nitrophenyl)-5-trifluoromethylthiazole-2-yl,4-(4-dimethylamino-3-trifluoromethylphenyl)-5-trifluoromethylthiazole-2-yl,4-(3-hydroxycarbonylphenyl)thiazole-2-yl,4-(4-hydroxycarbonylphenyl)thiazole-2-yl,4-(2,5-difluoro-4-(N-pyrrolidinyl)phenyl)thiazole-2-yl,4-(4-diethylamino-3-fluorophenyl)-5-fluorothiazole-2-yl,4-(4-aminophenyl)-5-trifluoromethylthiazole-2-yl,4-(4-(N-pyrrolidino)phenyl)thiazole-2-yl,4-(2,5-difluoro-4-(N-ethylpiperazine-N′-yl)phenyl)thiazole-2-yl,4-(4-diethylaminophenyl)-5-trifluoromethylthiazole-2-yl,4-(3,5-difluoro-4-diethylaminophenyl)-5-fluorothiazole-2-yl,4-(4-(N-morpholinyl)-2,5-difluorophenyl)thiazole-2-yl,4-(4-(N-piperidinyl)-2,5-difluorophenyl)thiazole-2-yl,4-(4-(N-pyrazolyl)-3-fluorophenyl)thiazole-2-yl,4-(4-chlorophenyl)-5-methylthiazole-2-yl, 5-methylthiazole-2-ylphenyl,4-(1-ethoxynaphth-3-yl)thiazole-2-yl,3-(4-chlorophenyl)isothiazole-5-yl, 4-(benzofuran-5-yl)thiazole-2-yl,3-(4-chlorophenyl)isoxazole-5-yl,4-(3-(N-pyrrolidinocarbonyl)phenyl)thiazole-2-yl,4-(4-dimethylaminophenyl)-5-phenylthiazole-2-yl,4-(3-(N-morpholinocarbonyl)phenyl)thiazole-2-yl,4-(4-diethylaminophenyl)-5-trifluoroacetylthiazole-2-yl,4-(4-diethylamino-3-fluorophenyl)-5-trifluoromethylthiazole-2-yl,4-(4-ethylamino-3-fluorophenyl)-5-trifluoromethylthiazole-2-yl,4-(4-diethylamino-3-fluoro-5-trifluoromethylphenyl)-5-trifluoromethylthiazole-2-yl,4-(4-(2-dimethylaminoethoxy)-3-fluorophenyl)thiazole-2-yl,4-(4-N-(pyrrolindinyl)phenyl)-5-ethylthiazole-2-yl,2-(4-(N-pyrrolidinyl)-3-fluorophenyl)pyridine-6-yl,4-(4-fluoro-3-(N-pyrrolidinyl)phenylthiazole-2-yl,4-(3-(N-pyrrolidinyl)-4-diethylaminophenyl)thiazole-2-yl,4-(4-(N-pyrrolyl)phenyl)-5-trifluoromethylthiazole-2-yl,2-(4-chlorophenyl)thiophene-5-yl, 4-(1-ethoxynaphth-5-yl)thiazole-2-yl,1-(4-chlorophenyl)pyrazole-3-yl,4-(4-(N-pyrrolidinyl)-2,5-difluorophenyl)-5-ethylthiazole-2-yl,4-(4-(N-pyrrolidinyl)-3-trifluoromethylphenyl)-5-methylthiazole-2-yl,4-(4-(N-piperidinyl)-3-trifluoromethylphenyl)-5-methylthiazole-2-yl,4-(4-chlorophenyl)-5-n-propylthiazole-2-yl,4-(4-ethoxy-2-(N-pyrrolidinyl)phenyl)thiazole-2-yl,4-(4-chloro-2-(N-pyrrolidinyl)phenyl)thiazole-2-yl,4-(4-(N-pyrrolidinyl)phenyl)-5-fluorothiazole-2-yl,4-(2-chloropyridine-5-yl)thiazole2-yl,2-(4-(N-pyrrolidinyl)phenyl)pyridine-6-yl,4-(4-(N-pyrrolidinyl)-3-fluorophenyl)-5-ethylthiazole-2-yl,4-(2-chloropyrid-3-yl)thiazole-2-yl,4-(4-(2-methyl-N-pyrrolidinyl)-3-fluorophenyl)thiazole-2-yl,4-(4-(N-pyrrolidinyl)-3-trifluoromethylphenyl)-5-methylthiazole-2-yl,4-(4-diethylamino-2-chlorophenyl)-5-ethylthiazole-2-yl,5-(4-chlorophenyl)-4-ethylthiazole-2-yl,4-(3,4-dichlorophenyl)-5-aminothiazole-2-yl,4-(6-chloroquinoline-3-yl)thiazole-2-yl,4-(2,4-bis(N-pyrrolidinyl)phenyl)thiazole-2-yl,4-(4-(2,4-dimethylpyrrolidine-N-yl)phenyl-3-fluorothiazole-2-yl,4-(thionaphthene-3-yl)thiazole-2-yl,4-(4-diethylaminophenyl)-5-ethylthiazole-2-yl,4-(2,6-diethoxy-3-bromophenyl)thiazole-2-yl, 2-thiophene-2-ylphenyl,2-(pyrrolidone-N-yl)phenyl, 1-phenyl-3-tert-butylpyrazole-5-yl,4-(4-(1-ethyltetrazolone-4-yl)phenylthiazole-2-yl,4-(4-diethylaminocarbonylphenyl)-5-phenylthiazole-2-yl,4-(4-(N-pyrrolidinyl)phenyl)-5-trifluoromethylthiazole-2-yl,4-(4-pyrimidine-5-ylphenyl)thiazole-2-yl,4-(4-morpholine-N-yl-3-fluorophenyl)-5-ethylthiazole-2-yl,4-(4-morpholine-N-yl-3-fluorophenyl)-5-propylthiazole-2-yl,4-(2-fluoropyridine-4-yl)thiazole-2-yl,4-(2-fluoropyridine-3-yl)thiazole-2-yl,4-(4-azacyclohept-N-yl-3-fluorophenyl)thiazole-2-yl,4-(2-chlororopyridine-4-yl)thiazole-2-yl,4-(2-methoxypyridine-5-yl)thiazole-2-yl,4-(3-fluoropyridine-4-yl)thiazole-2-yl,4-(3,4-dichlorophenyl)-5-nitrothiazole-2-yl,4-(1-methyl-2,3-dihydroindene-6-yl)thiazole-2-yl,4-(8-chloroquinoline-3-yl)thiazole-2-yl,4-(4-pyrrolidine-N-yl-2,3-difluorophenyl)-5-ethylthiazole-2-yl,4-(2-(pyrrolidine-N-yl)thiazole-4-yl)thiazole-2-yl,4-(4,5-benzo-2,1,3-oxadiazole-5-yl)thiazole-2-yl,4-(2-dimethlaminopyridine-5-yl)thiazole-2-yl,4-(2-fluoropyridine-5-yl)thiazole-2-yl,4-((4-methylpiperazine-N-yl)pyridine-6-yl)thiazole-2-yl,4-(3-fluoro-2-methylpyridine-6-yl)thiazole-2-yl, or wherein R¹ and R²are both methyl, D, R³, R⁴ and R⁵ are all H, n is 2 and Ar-Cy is3-(2-methylpyrimidine-4-yl)phenyl or wherein R¹ and R² are both methyl,R³, R⁴ and R⁵ are all H, D is trifluoromethyl, n is 2 and Ar-Cy is4-(4-chlorophenyl)thiazole-2-yl.d) Any one or more of the compounds described in WO2009/152261 A1;e) Any one or more compounds of structure I wherein R¹ and R² are bothmethyl, D, R³, R⁴ and R⁵ are all H, n is 1 and Ar-Cy is any one of4-phenylthiazole-2-yl, 4-(4-bromophenyl)thiazole-2-yl, 2- or4-(N-morpholinyl)phenyl, 4-(N-piperidinyl)phenyl,1-phenyl-3-methylpyrazole-5-yl, 4-(3,4-dimethylphenyl)thiazole-2-yl,4-(4-ethylphenyl)thiazole-2-yl, 4-(2,4,6-trimethylphenyl)thiazole-2-yl,4-(4-n-propylphenyl)thiazole-2-yl,4-(4-chlorophenyl)-5-methylthiazole-2-yl, 4-phenylphenyl,2-(N-pyrrolidinyl)phenyl, 4-(N-azacycloheptyl)phenyl,4-phenyl-5-ethylthiazole-2-yl, 2- or 4-cyclopentylphenyl, or wherein R²is methyl, R¹, D, R³, R⁴ and R⁵ are all H, n is 1 and Ar-Cy is4-cyclopentylphenyl.f) Any one or more of the compounds described in WO2007/073505 A1;g) Any one or more compounds of structure I wherein R¹ and R² are bothmethyl, D, R³, R⁴ and R⁵ are all H, n is 1 and Ar-Cy is any one of4-phenylthiazole-2-yl, 4-(4-bromophenyl)thiazole-2-yl, 2- or4-(N-morpholinyl)phenyl, 4-(N-piperidinyl)phenyl,1-phenyl-3-methylpyrazole-5-yl, 2-(2-benzimidazolyl)phenyl,4-(3,4-dimethylphenyl)thiazole-2-yl, 4-(4-ethylphenyl)thiazole-2-yl,4-(2,4,6-trimethylphenyl)thiazole-2-yl,4-(4-n-propylphenyl)thiazole-2-yl,4-(4-chlorophenyl)-5-methylthiazole-2-yl, 4-phenylphenyl,2-(N-pyrrolidinyl)phenyl, 4-(N-azacycloheptyl)phenyl,4-phenyl-5-ethylthiazole-2-yl,h) Any one or more of the compounds described in WO2009/002933 A1;i) A compound of structure I wherein R¹ and R² are both methyl, D, R³,R⁴ and R⁵ are all H, n is 1 and Ar-Cy is4-(4-diethylaminophenyl)thiazole-2-yl.j) Any one or more of the compounds described in WO2009/140519 A1;k) a compound of the formula shown below, or a salt thereof:

wherein, each of R¹ and R² is independently H, C1-C₆ alkyl, C₂-C₆alkenyl, or C₂-C₆ alkynyl, each of which is optionally substituted with1-4 R⁵; L is NR⁶SO₂, SO₂NR⁶, OC(O)NR⁶, NR⁶C(O)O, NR⁶C(O)NR⁶, NR⁶C(O),C(O)NR⁶, O, C(O), S, S(O), S(O)₂, NR₆, or CH₂, each of R^(a) and R isindependently cyclyl, heterocyclyl, aryl, heteroaryl, each of which isoptionally substituted with 1-4 R⁷; each of R¹ and R¹² is,independently, H, C₁-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆ alkynyl, halo,hydroxyl, alkoxy, amino, akylamino, thiol, alkylthiol, nitro, or cyano,each of which is optionally substituted with 1-2 R; each R isindependently halo, hydroxyl, alkoxy, amino, alkylamino, dialkylamino,cyano, nitro, amido, alkylamido, dialkylamido, thioyl, sulfonyl, cyclyl,heterocyclyl, aryl, or heteroaryl; each R⁶ is independently H, C₁-C₆alkyl, C₁-C₆ alkenyl, hydroxyC₁-C₆ alkyl, alkoxyC₁-C₆ alkyl, cyanoalkyl,haloalkyl, arylalkyl, S(O)alkyl, acyl, amino, amidyl, or S(O)₂H, aryl,alkoxyaryl; each R⁷ is independently C₁-C₆ alkyl, C₂-C₆ alkenyl, orC₂-C₆ alkynyl, halo, hydroxyl, alkoxy, oxo, aryl, heteroaryl, cyclyl,heterocyclyl, arylalkyl, heteroarylalkyl, cyclylalkyl,heterocyclylalkyl, aryloxy, arylalkoxy, amino, akylamino, dialkylamino,thioyl, alkylthioyl, sulfonyl, sulfonamidyl, amido, hydroxyl alkoxyl,alkoxy —C(O)OH, —C(O)Oalkyl, urea, sulfonylurea, acyl, nitro, cyano,each of which is optionally substituted with 1-3 R; each R⁸ isindependently C₁-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆ alkynyl, aryl,heteroaryl, cyclyl, halo, hydroxyl, alkoxy, oxo, aryloxy, amino,akylamino, dialkylamino, C(O)OH, —C(O)Oalkyl, thioyl, sulfonyl,sulfonamidyl, amido, urea, sulfonylurea, acyl, nitro, cyano, cyclyl,heterocyclyl, aryl, or heteroaryl; R⁹ is H, C₁-C₆ alkyl, C₂-C₆ alkenyl,or C₂-C₆ alkynyl, halo, C₁-C₆ haloalkyl, hydroxyl, alkoxy, aryloxy,arylalkoxy, amino, akylamino, dialkylamino, thioyl, alkylthioyl,sulfonyl, sulfonamidyl, amido, urea, sulfonylurea, acyl, nitro, cyano,each of which is optionally substituted with 1-3 R⁸; and m is 1, 2, 3,4, 5, or 6.l) Any one or more compounds of structure I wherein R¹ and R² are bothmethyl, D, R³, R⁴ and R⁵ are all H, n is 1 and Ar-Cy is any one of4-(4-bromophenyl)-6-trifluoromethylpyrimidine-2-yl,4-(3,4-dimethoxyphenyl)thiazole-2-yl,4-(2,4-dimethylphenyl)thiazole-2-yl, 4-(2-fluorophenyl)thiazole-2-yl,4-(3,4-difluorophenyl)-5-methylthiazole-2-yl,4-(3,4-methylenedioxyphenyl)thiazole-2-yl,4-(4-methoxynaphth-1-yl)thiazole-2-yl,4-(benzo-2-piperidino)thiazole-2-yl,4-(3-chloro-4-methoxyphenyl)thiazole-2-yl,4-(4-ethoxyphenyl)-5-methylthiazole-2-yl,4-(4-methylcarbonylaminophenyl)thiazole-2-yl,4-(4-bromophenyl)-5-methylthiazole-2-yl,4-(4-(2-methoxyethyl)phenyl)thiazole-2-yl,4-(4-methyl-3-nitrophenyl)thiazole-2-yl,4-(4-t-butylcarbonylaminophenyl)thiazole-2-yl,4-phenyl-5-(1-methylimidazole-2-yl)thiazole-2-yl,4-(4-(difluoromethoxy)phenyl)-5-methylthiazole-2-yl,4-(2,5-diethoxyphenyl)thiazole-2-yl,4-(4-fluorophenyl)-5-methylthiazole-2-yl,4-phenyl-5-methylcarbonylthiazole-2-yl,4-(3-chloro-4-fluorophenyl)thiazole-2-yl,4-(3-fluorophenyl)thiazole-2-yl, 4-(3-methoxyphenyl)thiazole-2-yl,4-(2-ethoxyphenyl)thiazole-2-yl, 4-(3-chlorophenyl)thiazole-2-yl,4-(2-methoxyphenyl)thiazole-2-yl, 4-(2-bromophenyl)thiazole-2-yl,4-(4-(dimethylamino)phenyl)thiazole-2-yl,4-(2-chlorophenyl)thiazole-2-yl, 4-(4-methoxyphenyl)thiazole-2-yl,4-(3-bromophenyl)thiazole-2-yl,4-(4-methylcarbonylamino-2-fluorophenyl)thiazole-2-yl,4-(4-isopropoxyphenyl)thiazole-2-yl,4-(2,3,4-trichlorophenyl)thiazole-2-yl,4-(3-chloro-4-methoxyphenyl)thiazole-2-yl,4-(3-fluoro-4-chlorophenyl)thiazole-2-yl,4-(3-ethoxyphenyl)thiazole-2-yl, 4-(3-nitrophenyl)thiazole-2-yl,4-(4-phenylphenyl)thiazole-2-yl, 4-(2,4-dimethoxyphenyl)thiazole-2-yl,4-(2,4-dimethylphenyl)thiazole-2-yl, 4-(4-cyanophenyl)thiazole-2-yl,4-(4-ethylphenyl)thiazole-2-yl, 4-(2-methylphenyl)thiazole-2-yl,4-(2-bromo-5-ethoxyphenyl)thiazole-2-yl,4-(2-chloro-4-ethoxyphenyl)thiazole-2-yl,4-(2-chloro-4-ethoxy-5-bromophenyl)thiazole-2-yl,4-(2-pyridyl)thiazole-2-yl, 4-(4-(2-hydroxylethoxy)phenyl)thiazole-2-yl,4-(3-(2-hydroxylethoxy)phenyl)thiazole-2-yl,4-(2,6-dichlorophenyl)thiazole-2-yl, 4-(3-methylphenyl)thiazole-2-yl,4,5-bis(4-methylphenyl)oxazole-2-yl, 4,5-diphenyloxazole-2-yl,4-(4-pyridyl)thiazole-2-yl, 4-(2-(2-hydroxyethoxy)phenyl)thiazole-2-yl,4-(3,4-dichlorophenyl)-5-methylthiazole-2-yl,4-(2,4,6-trichlorophenyl)thiazole-2-yl,4-(4-ethoxyphenyl)-5-phenylthiazole-2-yl,4-(4-chloro-3-ethoxyphenyl)thiazole-2-yl,4-(3,4,5-trichlorophenyl)thiazole-2-yl, 2-(4-pyridyl)1,3,4-thiadiazole-5-yl,4-(2-(2-hydroxyethoxy)-5-bromophenyl)thiazole-2-yl,3-(benzoxazole-2-yl)-2-methylphenyl,4-(2,3-dichlorophenyl)-5-ethylthiazole-2-yl,4-(3-trifluoromethylphenyl)thiazole-2-yl,4-(3,4-dichlorophenyl)-5-phenylthiazole-2-yl,4-(3,4-dichlorophenyl)-5-(4-methylphenyl)thiazole-2-yl,1-phenyl-3-(4-methylphenyl)pyrazole-5-yl,3-(4-fluorophenyl)pyrazole-5-yl,4,5-bis(3,4-dichlorophenyl)thiazole-2-yl; 5-phenylthiazole-2-yl,5-(4-chlorophenyl)thiazole-2-yl,4-(4-trifluoromethylphenyl)thiazole-2-yl,4-(4-methylsulfonylphenyl)thiazole-2-yl, 4,5-bis(2-furyl)thiazole-2-yl,4-(4-chlorophenyl)oxazole-2-yl,4-(4-trifluoromethoxyphenyl)thiazole-2-yl,3-(4-chlorophenyl)pyrazole-2-yl,2-(4-chlorophenyl)-1,3,4-oxadiazole-5-yl, 3-phenylpyrazole-5-yl,5-(4-methylphenyl)thiazole-2-yl,4-(4-chlorophenyl)-5-ethylthiazole-2-yl, 4-(4-nitrophenyl)thiazole-2-yl,4-(4-aminophenyl)thiazole-2-yl,4-(4-ethoxylphenyl)-5-methylthiazole-2-yl,4-(4-diethylamino-3-fluorophenyl)thiazole-2-yl,3-(thiophene-2-yl)pyrazole-5-yl,4-(4-chlorophenyl)-5-fluorothiazole-2-yl,4-(4-chlorophenyl)-5-aminothiazole-2-yl, 2-(pyrrole-N-yl)phenyl,2-(indole-2-yl)phenyl, 4-(benzimidazoloneyl)thiazole-2-yl,2-chlorophenylpyrid-4-yl, 3-(4-chlorophenyl)phenyl,4-(4-diethylamino-3-chlorophenyl)thiazole-2-yl,5-(4-ethoxyphenyl)thiazole-2-yl, 3-(4-chlorophenyl)-5-pyridyl,4-(3,4-dichlorophenyl)-5-isopropylthiazole-2-yl,4-(4-diethylamino-2-fluorophenyl)thiazole-2-yl,4-(4-(N-pyrrolidinyl)-3-bromophenyl)thiazole-2-yl,4-(4-fluoro-3-N-morpholinylphenyl)thiazole-2-yl,2-(4-chlorophenyl)pyrazine-6-yl, 4-(2,4,5-trichlorophenyl)thiazole-2-yl,5-(4-diethylaminophenyl)thiazole-2-yl,4-(1-ethoxynaphth-4-yl)thiazole-2-yl,4-(2,3-dihydrobenzofuran-5-yl)thiazole-2-yl,2-(3,4-dichlorophenyl)thiazole-4-yl,4-phenyl-5-(4-ethoxyphenyl)imidazole-2-yl,4-(4-(N-piperidinyl)phenyl)thiazole-2-yl,4-(3-diethylaminophenyl)thiazole-2-yl, 2-(benzothiazole-2-yl)phenyl,4-(3-(N-piperidinyl)phenyl)thiazle-2-yl,4-(3-dimethylaminophenyl)thiazole-2-yl, 4-(4-chlorophenyl)pyridine-2-yl,3-(4-diethylaminophenyl)phenyl, 4-(4-ethoxyphenyl)pyrazole-2-yl,4-(4-ethoxyphenyl)-5-methylpyrazole-2-yl,4(N-acetyl-2,3-dihydroindole-5-yl)thiazole-2-yl,4-(3,4-dichlorophenyl)pyrimidine-2-yl, 3-(thiazole-2-yl)phenyl,4-(quinoline-2-yl)thiazole-2-yl,4-(4-diethylamino-3,5-difluorophenyl)thiazole-2-yl,4-(3-(N-pyrrolidinyl)phenyl)thiazole-2-yl,4-(4-aminocarbonylaminophenyl)thiazole-2-yl,4-(4-chlorophenyl)-5-methoxylthiazole-2-yl,4-(4-(diethylaminosulfonyl)phenyl)thiazole-2-yl),4-(4-(dimethylaminosulfonyl)phenyl)thiazole-2-yl),4-(dihydrobenzoxazine-3-one-6-yl)thiazole-2-yl,4-(3,4-dichlorophenyl)imidazole-2-yl, 4-14,(4-ethoxyphenyl)oxazole-2-yl, 4-(4-chlorophenyl)oxazole-2-yl,3-(4-ethylthiazole-2-yl)phenyl, 3-(5-ethylthiazole-2-yl)phenyl,3-(4-methylthiazole-2-yl)phenyl,4-(4-diethylamino-2-chlorophenyl)thiazole-2-yl, 4-(4-diethylamino-2,5-difluorophenyl)thiazole-2-yl,4-(3-trifluoromethoxyphenyl)thiazole-2-yl, 2-(4-chlorophenyl)pyrid-6-yl,3-(4-diethylamino-3-fluorophenyl)phenyl,4-(4-diethylamino-2,3-difluorophenyl)thiazole-2-yl,4-(4-(N-piperidinylsulfonyl)phenyl)thiazole-2-yl,4-(N-acetylindole-5-yl)thiazole-2-yl,4-(2-ethoxynaphth-1-yl)thiazole-2-yl,1-(phenylmethyl)-4-(3,4-diphenyl)imidazole-2-yl,4-(N-ethyl-2,3-dihydroindole-5-yl)thiazole-2-yl, 3-(4-chlorophenyl)1,2,4-thiadiazole-5-yl,4-(N-acetyl-2,3-dihydroindole-6-yl)thiazole-2-yl,4-(N-ethylindole-5-yl)thiazole-2-yl, 3-(4-chlorophenyl)thiophene-5-yl,1-(4-chlorophenyl) 1,2,4-thiazole-3-yl, 1-phenylpyrazole-4-yl,4-(N-acetylindole-6-yl)thiazole-2-yl,4-N-methyl-2,3-dihydroindole-5-yl)thiazole-2-yl,4-(4-diethylamino-2-chloro-5-fluorophenyl)thiazole-2-yl,4-(3-dimethylaminosulfonylphenyl)thiazole-2-yl,4-(3-methylsulfonylaminophenyl)thiazole-2-yl,4-(3-diethylaminosulfonylphenyl)thiazole-2-yl,2-(4-chlorophenyl-1,3,5-triazine-4-yl,4-(4-(miorpholin-N-ylsulfonyl)phenyl)thiazole-2-yl),4-(4-(N-pyrrolidinosulfonyl)phenylthiazole-2-yl),4-(4-chloro-3-diethylaminophenyl)thiazole-2-yl,4-(4-chloro-3-trifluoromethylphenyl)thiazole-2-yl,1-ethyl-4-(2,3-dichlorophenyl)imidazole-2-yl,1-methyl-4-(2,3-dichlorophenyl)imidazole-2-yl, 2-(1-naphthyl)5-pyridyl,2-(3,4-dichlorophenyl)5-pyridyl,4-(4-chlorophenyl)-5-dimethylaminomethylthiazole-2-yl,4-(3-diethylaminophenyl)-5-methylthiazole-2-yl,4-(4-diethylamino-3-fluorophenyl)-5-ethylthiazole-2-yl,4-(4-chlorophenyl)-5-(2-dimethylaminoethyl)thiazole-2-yl,4-(4-trifluoromethyl-2-fluorophenyl)thiazole-2-yl,4-(3-trifluoromethyl-5-fluorophenyl)thiazole-2-yl,4-(4-trifluoromethyl-3-fluorophenyl)thiazole-2-yl,4-(3-trifluoromethyl-4-fluorophenyl)thiazole-2-yl,4-(3,4-dichlorophenyl)-5-methoxycarbonylthiazole-2-yl,4-(1-bromo-2-ethoxynaphth-6-yl)thiazole-2-yl,4-(3,4-dichlorophenyl)-5-methoxycarbonylmethylthiazole-2-yl,1,5-dimethyl-4-(3,4-dichlorophenyl)imidazole-2-yl,N-(4-chlorophenyl)pyrrole-3-yl,4-(3,4-dichlorophenyl)-5-aminocarbonylthiazole-2-yl,4-(2-ethoxynaphth-6-yl)thiazole-2-yl,4-(3,4-dichlorophenyl)-5-hydroxycarbonylmethylthiazole-2-yl,4-(4-diethylamino-2,5-difluorophenyl)-5-ethylthiazole-2-yl,4-(4-(N-pyrrolidinyl)-3-fluorophenyl)thiazole-2-yl,4-(4-diethylamino-3-ethoxyphenyl)thiazole-2-yl,4-(4-diethylamino-2-ethoxyphenyl)thiazole-2-yl,4-(4-(N-pyrrolidinyl)-3-(trifluoromethyl)phenyl)thiazole-2-yl,4-(4-diethylamino-3-trifluoromethylphenyl)thiazole-2-yl,4-(1-n-butyl-4-(3,4-dichlorophenyl)imidazole-2-yl,1-(4-chlorophenyl)imidazole-4-yl, 4-(2-indolyl)thiazole-2-yl,5-(4-chlorophenyl)isoxazole-2-yl, 5-(4-chlorophenyl)1,2,4-oxadiazole-3-yl,1-ethyl-5-methyl-4-(3,4-dichlorophenyl)imidazole-2-yl,4-phenyl-5-(4-ethoxyphenyl)oxazole-2-yl, 2-naphthyl-5-pyridyl,2-(4-chlorophenyl)oxazole-5-yl,4-(3-methylethylaminophenyl)-5-ethylthiazole-2-yl,4-(4-ethoxyl-3-diethylaminophenyl)thiazole-2-yl,4-(4-diethylaminophenyl)-5-fluorothiazole-2-yl,4-(4-(4-ethylpiperazine-1-yl)-3-trifluoromethylphenyl)thiazole-2-yl,4-(3-ethoxycarbonylphenyl)thiazole-2-yl,4-(4-(N-pyrrolidinyl)-3,4-difluorophenyl)thiazole-2-yl,4-(4-chlorophenyl)-5-trifluoromethylthiazole-2-yl,4-(4-diethylamino-3-cyanophenyl)thiazole-2-yl,4-(2-ethoxynaphth-5-yl)thiazole-2-yl,4-(4-chlorophenyl)isothiazole-3-yl,4-(N-methylindole-5-yl)thiazole-2-yl,4-(3,4-dichorophenyl)-5-(2-hydroxyethyl)thiazole-2-yl,4-(4-chlorophenyl)thiazole-2-yl in which case D is trifluoromethyl,1-(4-chlorophenyl) 1,2,3-triazole-4-yl,4-(4-(2H-imidazole-1-yl)phenyl)thiazole-2-yl,4-(4-tetrazole-5-ylphenyl)thiazole-2-yl,4-(4-methyl-4-methoxyphenylmethylaminophenyl)thiazole-2-yl,4-(4-(N-pyrrolidinyl)-3,5-difluorophenyl)-5-fluorothiazole-2-yl,4-(4-N-morpholinyl-3-trifluoromethylphenyl)thiazole-2-yl,4-(4-(N-piperidinyl)-3-(trifluoromethyl)phenyl)thiazole-2-yl,4-(4-nitrophenyl)-5-trifluoromethylthiazole-2-yl,4-(4-diethylamino-3-trifluoromethylphenyl)-5-trifluoromethylthiazole-2-yl,4-(3-hydroxycarbonylphenyl)thiazole-2-yl,4-(4-hydroxycarbonylphenyl)thiazole-2-yl,4-(4-N-pyrrolidinyl-25-difluorophenyl)thiazole-2-yl,4-(4-diethylamino-3-fluorophenyl)-5-fluorothiazole-2-yl,4-(4-diethylamino-3-trifluoromethylphenyl)-5-trifluoromethylthiazole-2-yl,4-(4-aminophenyl)-5-trifluoromethylthiazole-2-yl,4-(4-(N-pyrrolidinyl)phenyl)thiazole-2-yl,4-(4-(4-ethylpiperazine-1-yl)phenyl-2,5-difluorophenyl)thiazole-2-yl,4-(4-diethylaminophenyl)-5-trifluoromethylthiazole-2-yl,4-(4-diethylamino-3,5-difluorophenyl)-5-fluorothiazole-2-yl,4-(4-(N-morpholinyl)-2, 5-difluorophenyl)thiazole-2-yl,4-(4-(N-piperidinyl)-2,5-difluorophenyl)thiazole-2-yl,4-(4-(N-pyrazolyl)-3-fluorophenyl)thiazole-2-yl,4-(4-chlorophenyl)-5-methylthiazole-2-yl,3-(5-methylthiazole-2-yl)phenyl, 4-(1-ethoxylnaphth-3-yl)thiazole-2-yl,5-(4-chlorophenyl)isothiazole-3-yl, 4-(5-benzofuryl)thiazole-2-yl,3-(4-chlorophenyl)isoxazole-5-yl,4-(3-N-pyrrolidinylcarbonylphenyl)thiazole-2-yl,4-(4-diethylaminophenyl)-5-phenylthiazole-2-yl,4-(3-N-morpholinylcarbonylphenyl)thiazole-2-yl,4-(4-diethylaminophenyl)-5-trifluoromethylcarbonylthiazole-2-yl,4-(4-diethylamino-3-fluorophenyl)-5-trifluoromethylthiazole-2-yl,4-(4-ethylamino-3-fluorophenyl)-5-trifluoromethylthiazole-2-yl,4-(4-diethylamino-3-fluoro-5-trifluoromethylphenyl)-5-trifluoromethylthiazole-2-yl,4-(4-(2-dimethylaminoethoxy)-3-fluorophenyl)thiazole-2-yl,4-(4-(N-pyrrolidinyl)phenyl)-5-ethylthiazole-2-yl,2-(4-(N-pyrrolidinyl)-3-fluorophenyl)pyridin-5-yl,4-(4-fluoro-3-9N-pyrrolidinyl0phenyl)thiazole-2-yl, 4-(4-diethylamino-3-(N-pyrrolidinyl)phenyl)thiazole-2-yl,4-(4-(N-pyrrolyl)phenyl)-5-(trifluoromethyl)thiazole-2-yl,2-(4-chlorophenyl)thiophene-5-yl, 4-(1-ethoxynaphth-5-yl)thiazole-2-yl,1-(4-chlorophenyl)pyrazole-3-yl,4-(4-(N-pyrrolidinyl)-2,5-difluorophenyl)-5-ethylthiazole-2-yl,4-(4-(N-pyrrolidinyl)-3-trifluoromethylphenyl)-5-methylthiazole-2-yl,4-(4-(N-piperidinyl)-3-trifluoromethylphenyl)-5-methylthiazole-2-yl,4-4(4-chlorophenyl)-5-n-propylthiazole-2-yl,4-(4-ethoxyl-2-(N-pyrrolidinyl)phenyl)thiazole-2-yl,4-(4-chloro-2-(N-pyrrolidinyl)phenyl)thiazole-2-yl,4-(4-(N-pyrrolidinyl)phenyl)-5-fluorothiazole-2-yl,4-(2-chloropyridine-5-yl)thiazole-2-yl,2-(4-(N-pyrrolidinyl)phenyl)pyridine-5-yl,4-(4-(N-pyrrolidinyl)-3-fluorophenyl)-5-ethylthiazole-2-yl,4-(2-chloropyridine-3-yl)thiazole-2-yl,4-(4-(2-methylpyrrolidine-1-yl)-3-fluorophenyl)thiazole-2-yl,4-(4-(N-pyrrolidinyl)-3-(trifluoromethyl)phenyl)-5-n-propylthiazole-2-yl,4-(4-diethylamino-2-chlorophenyl)-5-ethylthiazole-2-yl,4-ethyl-5(4-chlorophenyl)thiazole-2-yl,4-4(3,4-dichlorophenyl)-5-aminothiazole-2-yl,4-(6-chloroquinoline-3-yl)thiazole-2-yl,4-(2,4-bis(N-pyrrolidinyl)phenyl)thiazole-2-yl,4-(4-(2,5-dimethylpyrrolidine-1-yl)³-fluorophenyl)thiazole-2-yl,4-benzothiophene-3-ylthiazole-2-yl,4-(4-diethylaminophenyl)-5-ethylthiazole-2-yl,4-(4-methylaminophenyl)thiazole-2-yl,4-(2,6-diethoxy-3-bromophenyl)thiazole-2-yl, 2-(thiophene-2-yl)phenyl,2-(pyrrolidone-N-yl)phenyl, 1-phenyl-3-t-butylpyrazole-5-yl,4-(4-(1-ethyltetrazolone-4-yl)phenyl)thiazole-2-yl,4-(4-diethylaminocarbonylphenyl)-5-phenylthiazole-2-yl,4-(4-(N-pyrrolyl)phenyl)-5-(trifluoromethyl)thiazole-2-yl,4-(pyrimidine-5-yl)thiazole-2-yl,4-(4-(N-morpholinyl)-3-fluorophenyl)-5-n-propylthiazole-2-yl,4-(2-fluoropyrine-4-yl)thiazole-2-yl,4-(2-fluoropyrine-3-yl)thiazole-2-yl,4-(4-(N-azacycloheptyl)-3-fluorophenyl)thiazole-2-yl,4-(2-chloropyridine-4-yl)thiazole-2-yl,4-(2-methoxylpyridine-5-yl)thiazole-2-yl,4-(3-fluoropyrine-4-yl)thiazole-2-yl,4-(3,4-dichlorophenyl)-5-nitrothiazole-2-yl,4-(N-methyl-2,3-dihydroindole-6-yl)thiazole-2-yl,4-(8-chloroquinoline-3-yl)thiazole-2-yl,4-(4-(N-pyrrolidinyl)-2,3-difluorophenyl)-5-ethylthiazole-2-yl,4-(2-(N-pyrrolidinyl)thiazole-4-yl)thiazole-2-yl,4-(benzofurazan4-yl)thiazole-2-yl,4-(2-dimethylaminopyridine-5-yl)thiazole-2-yl,4-(2-fluoropyridine-5-yl)thiazole-2-yl,4-(2-(4-methylpiperidine-1-yl)pyridine-6-yl)thiazole-2-yl,4-(3-fluoro-2-methylpyridine-6-yl)thiazole-2-yl.m) Any one or more of the compounds described in WO2009/140517 A1;n) a compound of the formula below or a salt thereof,

wherein each of R¹ and R² is, independently, H, C₁-C₆ alkyl, C₂-C₆alkenyl, or C₂-C₆ alkynyl, each of which is optionally substituted with1-4 R⁵; L is NR⁶SO₂, SO₂NR⁶, C(O)NR⁶, NR⁶C(O), OC(O)NR⁶, NR⁶C(O)O,NR⁶C(O)NR⁶, S, S(O), S(O)₂, NR⁶, CH₂, O, C(O)NS(O)₂, S(O)₂NC(O),heteroaryl, or cyclyl; R is C₄-C₁₄ cyclyl, heterocyclyl, aryl, orheteroaryl, each of which is optionally substituted with 1-4 R⁷; each R⁵is independently halo, hydroxyl, alkoxy, thiol, alkylthio, amino,alkylamino, dialkylamino, cyano, nitro, amido, alkylamido, dialkylamido,thioyl, sulfonyl, cyclyl, heterocyclyl, aryl, or heteroaryl; each R⁶ isindependently H, C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl, aryl,arylalkyl, or acetyl; each R⁷ is independently C₁-C₆ alkyl, C₂-C₆alkenyl, C₂-C₆ alkynyl, cyclyl, heterocyclyl, aryl, heteroaryl, halo,hydroxyl, alkoxy, thiol, alkylthio, aryloxy, arylalkoxy, amino,akylamino, dialkylamino, thioyl, alkylthioyl, sulfonyl, sulfonamidyl,amido, urea, sulfonylurea, hydroxyl alkoxyl, alkoxy alkoxyl, acyl,nitro, or cyano, each of which is optionally substituted with 1-3 R⁸;each R⁸ is independently C₁-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆ alkynyl,halo, hydroxyl, alkoxy, thiol, alkylthio, aryloxy, amino, akylamino,dialkylamino, thioyl, sulfonyl, sulfonamidyl, amido, urea, sulfonylureaacyl, nitro, cyano, cyclyl, heterocyclyl, aryl, or heteroaryl; R⁹ isindependently H, C₁-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆ alkynyl, halo,hydroxyl, alkoxy, aryloxy, arylalkoxy, amino, akylamino, dialkylamino,thioyl, alkylthioyl, sulfonyl, sulfonamidyl, amido, urea, sulfonylurea,acyl, nitro, cyano, and is optionally substituted with 1-3 R⁸; each ofR^(n)-R¹⁴ is, independently, H, C₁-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆alkynyl, halo, hydroxyl, alkoxy, amino, akylamino, thiol, alkylthiol,nitro, or cyano, each of which is optionally substituted with 1-2 R⁸;each of m and n is, independently, 0, 1, 2, 3, 4, 5, or 6.o) Any one or more, optionally all, of the compounds 7, 13, 17, 27, 28,39, 42, 43, 44, 58, 65, 71 and 80, and also optionally 83, describedhereinafter.

The following particular options may be used in conjunction with thefirst aspect, either individually or in any suitable combination.

R¹ and R² may, independently, be methyl or ethyl. They may both bemethyl. R³ and R⁴ may both be H. They may both be alkyl, e.g. methyl orethyl. One of R³ and R⁴ may be alkyl (e.g. methyl or ethyl) and theother H. R⁵ may be hydrogen. It may be methyl or may be some otheralkyl.

If Ar is a 6-membered ring, it may be 1,4-disubstituted. If Ar is a5-membered ring it may be 1,3-disubstituted. Ar may be a ring which isnot 1,2-disubstituted. In this context, “disubstituted” refers tosubstitution by Cy and NR⁵.

Ar may be a disubstituted benzene ring (i.e. a phenylene ring), adisubstituted thiophene ring or a disubstituted nitrogen heterocyclehaving between 1 and 4 nitrogen atoms. It may be a 6 membered aromaticring having between 0 and 2 nitrogen atoms. It may be a ring that is nota 2-thiazolyl ring. It may be a pyridazine ring, e.g. apyridazin-3,6-diyl.

Cy may be a 5 or 6 membered aromatic ring having between 0 and 2nitrogen atoms, 0 or 1 sulfur atoms and 0 or 1 oxygen atoms.Alternatively it may be piperazine. The piperazine may be substituted onboth nitrogen atoms. Cy may contain no chlorine. It may be a group thatis not a chlorophenyl group (optionally additionally substituted). Insome embodiments compound I has no chlorine.

n may be 1.

D may be H.

The modulating may be inhibiting.

In a particular embodiment, R¹ and R² are both methyl, R³, R⁴, R⁵ and Dare all H and n is 1. In a specific instance of this embodiment, Ar is a6-membered ring having 1 or 2 nitrogen atoms and having no substituentsother than Cy and the amide nitrogen, these being in a 1,4-relationshipon the ring.

The compound may have an IC₅₀ against STF3A of less than 10 micromolar,or less than 5 micromolar or less than 1 micromolar, or less than 0.1micromolar.

The compound may not modulate, or may not inhibit, TRPA1. It may notinhibit TRPA1 at an IC₅₀ of <5 micromolar or at an IC₅₀ of <10micromolar.

The first aspect also includes all enantiomers and diastereomers of thecompound, as well as salts of the compounds. Suitable salts includepharmaceutically and/or veterinarially acceptable salts, for example thehydrochloride salts. The free bases of the compounds are alsoencompassed.

In a second aspect of the invention there is provided use of a compoundas defined in the first aspect for modulating, optionally inhibiting,Wnt activity and/or porcupine activity. There is also provided a methodof modulating, optionally inhibiting, Wnt activity (e.g. Wnt secretion)and/or porcupine activity comprising exposing cells, or a Wnt protein ora Wnt receptor, to a compound as defined in the first aspect. The cellsmay be cells that over-express Wnt protein. The method may be an invitro method or it may be an in vivo method. Without wishing to be boundby theory, the inventors hypothesise that the compounds defined in thefirst aspect inhibit the secretion of Wnt proteins. The compounds of theinvention are capable of inhibiting porcupine, which is essential andspecific for the palmitoylation of Wnt proteins before secretion. Thusin an embodiment there is provided a method of inhibiting Wnt secretionin a cell, said method comprising exposing said cell to a compound asdefined in the first aspect. In another embodiment there is provided amethod of inhibiting Wnt secretion in a cell which over-expresses Wntprotein, said method comprising exposing said cell to a compound asdefined in the first aspect.

In a third aspect of the invention there is provided use of a compoundas defined in the first aspect for treatment of a disease or conditionassociated with Wnt pathway activity. The Wnt pathway activity may beexcessive activity. This aspect also includes a method for treating saiddisease or condition, comprising administering to a subject in needthereof a therapeutically effective amount of the compound. The subjectmay be a human or may be a non-human, e.g. a non-human mammal or othernon-human animal.

The disease or condition may be selected from the group consisting ofcancer, fibrosis, stem cell and diabetic retinopathy. The cancer may bea cancer characterised by abnormal, optionally high, Wnt activity.

In a fourth aspect of the invention there is provided use of a compoundas defined in the first aspect for the manufacture of a medicament forthe treatment of a disease or condition associated with abnormal,optionally high, Wnt pathway activity. The disease or condition may beselected from the group consisting of cancer, fibrosis, stem cell anddiabetic retinopathy, rheumatoid arthritis, psoriasis and myocardialinfarction. There is also provided a composition or medicament for thetreatment of such a disease or condition, said composition or medicamentcomprising a compound as defined in the first aspect together with oneor more pharmaceutically acceptable carriers, diluents or adjuvants.

The disease or condition may be a cancer, such as cervical, colon,breast, bladder, head and neck, gastric, lung, ovarian, prostate,thyroid, non-small-cell lung, as well as chronic lymphocytic leukemia,mesothelioma, melanoma, pancreatic adenocarcinoma, basal cell carcinoma,osteosarcoma, hepatocellular carcinoma, Wilm's tumour ormedulloblastoma. The disease or condition may be a severe fibroticdisease, such as pulmonary fibrosis, liver fibrosis, skin fibrosis orrenal fibrosis. It may be a degenerative disease. It may be a metabolicdisease such as diabetic retinopathy.

In a fifth aspect of the invention there is provided a compound asdefined in the first aspect for use in therapy.

In a sixth aspect of the invention there is provided a pharmaceuticalcomposition comprising a compound according to the first aspect togetherwith one or more pharmaceutically acceptable carriers, diluents oradjuvants.

In a seventh aspect of the invention there is provided an anhydrous formof the free base of2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(6-phenylpyridazin-3-yl)acetamide.There is also provided a pharmaceutical composition comprising saidanhydrous free base, a method of treating or preventing a proliferativedisorder comprising administering a therapeutically effective amount ofsaid anhydrous free base to a subject in need thereof and use of saidanhydrous free base either in the treatment of a proliferative disorderor in the manufacture of a medicament for the treatment of aproliferative disorder.

BRIEF DESCRIPTION OF DRAWINGS

FIG. 1 shows tumour weights and volumes in mice treated with Compound 5:

FIG. 2 shows images of tumours treated with Compound 5.

FIG. 3 shows the time-course of p-LRP6 (Ser 1490) inhibition (in vitro)induced by porcupine inhibitors. It is a Western blot analysis of PA-1teratocarcinoma cells treated with 2 μM of Compound 5, for the timepoints indicated. As a control, cells treated with either growth mediumalone or the vehicle (DMSO) for 48 h were included. (A) Western blotswith antibodies as indicated on the left. (B) Densitometric analysis ofpLRP6 relative to total LRP6.

FIG. 4 shows pLRP6 in vitro inhibition-dose titration. It is a Westernblot analysis of HPAF-II pancreatic adenocarcinoma cells treated withthe indicated doses of Compound 5 for 6 h. The positive controls are acell lysate of untreated STF3A cells and cells treated with vehicle only(DMSO). (A) Western blots with antibodies as indicated on the left. (B)Densitometric analysis of pLRP6 relative to total LRP6.

DESCRIPTION OF EMBODIMENTS

The invention relates to the preparation and the use of new compoundsthat modulate Wnt activity, to methods of using the compounds, as asingle agent or in combination, for treating or preventing diseases andconditions associated with Wnt pathway activity, in particular having adysfunction linked to Wnt signalling pathway i.e. cancer, fibrosis, stemcell and diabetic retinopathy. Thus the invention relates to a class ofcompounds that act as modulators of the Wnt pathway and topharmaceutical compositions comprising these compounds and to their usefor the preparation of a medicament for the treatment of diseases havinga dysfunction linked to Wnt signalling pathway where Wnt plays a role inproliferation of cancer via multiple mechanisms, including a key role instem cell maintenance. Dysfunction of the Wnt pathway is related toconditions including, but not limited to, cancers such as cervical,colon, breast, bladder, head and neck, gastric, lung, ovarian, prostate,thyroid, non-small-cell lung, as well as chronic lymphocytic leukemia,mesothelioma, melanoma, pancreatic adenocarcinoma, basal cell carcinoma,osteosarcoma, hepatocellular carcinoma, Wilm's tumour andmedulloblastoma and other diseases with high Wnt expression such asfibrosis (including skin, idiopathic pulmonary, liver, renalinterstitial, myocardial, infarct and liver) and diabetic retinopathy.Respiratory conditions, or respiratory tumours, may in certainembodiments not be conditions treated by the present invention.

Many of the compounds of the present invention are1,3-dimethyl-3,7-dihydro-1H-purine-2,6-diones or1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurines and related compounds.The general structure of these is structure (I) as defined earlier. Inthis definition the following may apply.

Alkyl groups may be linear or may be branched. They may be C1 to C12 ormay be more than C12. They may for example be C1 to C6, C1 to C3, C3 toC6 or C6 to C12, e.g. methyl, ethyl, propyl, isopropyl, butyl, isobutyl,tertiary butyl, pentyl, neopentyl, hexyl, octyl, isooctyl, decyl ordodecyl. In certain instances they may contain cyclic structures. Thusthey may for example be, or may contain, cyclohexyl, methylcyclohexyl,isopropylcyclopentyl, cyclobutylethyl etc. In certain embodiments theyare not cage structures such as adamantyl.

Aryl groups may be homoaromatic. They may be benzenoid. They may bemonocyclic, bicyclic or polycyclic (i.e. contain more than 2 rings).They may comprise fused and/or unfused rings. They may, unless otherwisespecified, have any suitable substitution pattern, e.g. ortho, meta,para. Unless specified, they may have any appropriate number ofsubstituents (e.g. a monocyclic aromatic may have from 1 to 5substituents, a fused bicyclic aromatic may have from 1 to 7substituents etc.).

Heteroaryl groups may have 1 heteroatom, or may have 2, 3 or 4heteroatoms or in some cases more than 4. The heteroatom(s) are commonlyselected (independently) from N, S and O, however in some instancesother heteroatoms may be present. Heteroaryl groups typically have 5 or6 ring atoms unless they are bicyclic or polycyclic. Unless otherwisespecified, they may have any suitable substitution pattern and may haveany appropriate number of substituents. Heteroaryl groups may bemonocyclic or bicyclic or polycyclic. The fused rings may each be eithera heteroaryl ring or a homoaryl ring, provided that at least one isheteroaryl.

Non-aromatic rings may, unless otherwise specified, be carbocyclic ormay contain one or more heteroatoms, e.g. 1, 2, 3 or 4 heteroatoms. Eachheteroatom may, independently, be N, S or O, or some other heteroatom.The rings may have from 4 to 8 ring atoms, commonly 5 or 6.

Suitable examples include piperazinyl and morpholinyl rings. Group Cy instructure (I) may be, in some embodiments, an example of such rings.

The term “optionally substituted” signifies that one or moresubstituents may be present or there may be no substituents.Substituents may or may not be present on the above groups (alkyl, aryl,heteroaryl, non-aromatic rings). There may be 0, 1, 2, 3 or 4 or morethan 4 substituents on a group, as dictated by the structure of thegroup. Possible substituents include halogens (e.g. fluorine, chlorineor bromine), alkyl groups, alkoxy groups (i.e. O-alkyl, where alkyl isas defined above), aryloxy groups (i.e. O-aryl, where aryl is as definedabove), ester, amide or sulfonate ester groups (i.e. CO₂R, CONHR, SO₃R,where R is alkyl as defined above), however other substituents mayadditionally or alternatively be present. In cases where substituentsare shown, the term “optionally substituted” indicates the possibilityof additional substituents that are not shown. Thus for example instructure I, when it is stated that Ar is “optionally substituted”, thisindicates the possibility of further substituents additional to Cy andNR⁵, but does not indicate the possibility that either Cy or NR⁵ mightbe absent. Thus, for example, in cases where it is stated that Ar is a“disubstituted” aromatic ring, this should be taken- to mean that thereare only two substituents on the ring, i.e. no additional substituentsother than Cy and the amide nitrogen. For example, the 1,4-phenylenegroup in compound 1 is regarded as “disubstituted”.

In structures (I) and (II), n may be 1 to 5. It may be 1 to 3. It may beany one of 1, 2, 3, 4 or 5. Substituent D may be H, halogen, alkyl,cycloalkyl, aryl, or dialkylamino, each (other than H and halogen) beingoptionally substituted. Examples include hydrogen, chlorine, bromine,methyl, ethyl, propyl, cyclopropyl, phenyl, trifluoromethyl,dimethylamino, N-piperidinyl, N-piperazinyl, N-methyl-N′-piperazinyletc. In many embodiments, n is 1.

In some cases, the substituents R³ and R⁴ are the same, and in othersthey are different. In the event that they are different, they give riseto stereochemistry at the carbon atom to which they are attached. Ingeneral the stereochemistry at that carbon (or at each carbon) may be(S) or (R). In the particular example where n is 1 and one of R³ and R⁴is H and the other is an alkyl group, a preferred stereochemistry is asshown in structure (II), where Me represents the alkyl group. In caseswhere the alkyl group is methyl, this stereochemistry is particularlypreferred.

Examples of group Ar in structure (I) include 1,4-phenylene,2,5-pyridinediyl, 3,6-pyridazinediyl, 2,5-pyrazinediyl,2,5-thiophenediyl, 2,4-thiophenediyl, 2,5-furandiyl, 2,4-furandiyl, etc.Examples of group Cy in structure (I) include phenyl, thiazole-2-yl,thiophene-2-yl, thiophene-3-yl, pyridine-1-yl, pyridine-2-yl,pyridine-3-yl, pyridazine-3-yl, pyridazine-4-yl, pyrimidine-2-yl,pyrimidine-4-yl, pyrimidine-4-yl, N-imidazolyl, 2-, 4- or 5-thiazolyl,2-, 4- or 5-oxazolyl, N-morpholinyl or N′ substituted N-piperazinyl.Suitable substituents on the N′ position of the piperazinyl substituentinclude —C(═O)X, where X is t-butoxy, neopentyl, methyl, phenyl,p-chlorophenyl, benzyl, α,α-difluorobenzyl, chlorobenzyl, fluorobenzyletc.

In some embodiments, R¹ and R² are the same. They may be both methyl.They may be both ethyl. In the latter case, Ar may be 1,4-phenylene andCy may be thiophene-3-yl.

In some embodiments D is H. In other embodiments, D is methyl,cyclopropyl, trifluoromethyl, phenyl, dimethylamino, morpholin-N-yl,thiophene-3-yl or bromo. In the event that D is not H (e.g. is methyl,cyclopropyl, trifluoromethyl, phenyl, dimethylamino, morpholin-N-yl,thiophene-3-yl or bromo), Ar may be 1,4-phenylene and Cy may bethiophene-3-yl.

In some embodiments, n is 1 and in others it is 2 or 3. In the eventthat n is 2 or 3, Ar may be 1,4-phenylene and Cy may be thiophene-3-ylor thiazole-2-yl.

In some embodiments, R³ and R⁴ are either both H or both methyl. Inother embodiments, one is H and the other is methyl or ethyl. In theevent that they are not both H, Ar may be 1,4-phenylene. Alternatively,if they are not both H, either Ar is 1,4-phenylene or Cy is phenyl oreither is thiazole-2-yl.

In some embodiments R⁵ is H.

In some embodiments Ar is 1,4-phenylene and Cy is thiophene-3-yl orthiazole-2-yl. In particular embodiments Ar is 1,4-phenylene and Cy isthiophene-3-yl.

In a particular embodiment, R¹ and R² are both Me, D is H, n is 1 and R³and R⁴ are both H. In this embodiment, it is preferred that if Ar is1,4-phenylene, Cy is not thiophene-3-yl.

In a variation of this embodiment, either R¹ and R² are not both methyl,or D is not H, or n is not 1, or R³ and R⁴ are not both H (optionallymore than one, and in particular instances all, of these apply) and Aris 1,4-phenylene and Cy is thiophene-3-yl or thiazole-2-yl (optionallyAr is 1,4-phenylene and Cy is thiophene-3-yl).

In some embodiments, any one or more, optionally all, of compounds 7,13, 27. 28, 39, 42, 43, 44, 58, 65, 71, 80 and 83 as defined hereinaftermay be excluded from the scope of the invention. In some embodiments,any one or more, optionally all, of compounds 8, 12, 55 and 85 may alsobe excluded.

In some embodiments, Ar and Cy are not both optionally substitutedphenyl rings. In some embodiments, at least on of Ar and Cy isheteroaromatic or non-aromatic. In some embodiments at least one of Arand Cy is heteroaromatic.

In some embodiments, if Ar is 1,4-phenylene or 2,5-pyridyl, Cy has nomore than 1 ring nitrogen atom. In other embodiments, if Ar is a5-membered ring, it is not oxazolediyl. In yet other embodiments, if Aris a 5-membered ring, it may be thiophenediyl, e.g. thiophene-2,4-diyl.In the context of the present specification, reference to “if A then B”should be taken to indicate the possibilities either that A is not thecase or that both A and B are the case. Therefore for example, thestatement “if Ar is a 5-membered ring, it may be thiophenediyl” may betaken to mean that either Ar is not a 5-membered ring, or else Ar is athiophenediyl ring. In such instances, Ar could be for example, apyridinediyl ring, but could not be a furandiyl ring.

It will be understood that many (although not all) of the limitationsset out above in various embodiments may be used together incombination, and the present invention explicitly contemplates suchcombinations where they are practicable.

Specific (but non-limiting) examples of the compounds of the presentinvention are set out below:

The compounds of the present invention may have an IC₅₀ against STF3A ofless than about 10 micromolar. The IC₅₀ may be less than about 5, 2, 1,0.5, 0.2 or 0.1 micromolar. It may be between about 0.01 and about 10micromolar, or between about 0.01 and 5, 0.01 and 1, 0.01 and 0.5, 0.01and 0.1, 0.01 and 0.05, 0.1 and 5, 0.1 and 1, 0.1 and 0.5, 0.1 and 10,0.5 and 10, 1 and 10, 5 and 10, 1 and 5 or 0.1 and 0.5, e.g. about 0.01,0.02, 0.03, 0.04, 0.05, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1,2, 3, 4, 5, 6, 7, 8, 9 or 10 micromolar. A suitable method for testingIC₅₀ is as follows: approximately 5000 cells in 75 μl culture media areseeded in each well of black 96 well plates and incubated overnight at37° C. 25 μl of serially diluted compound is then added to the cellsgiving final concentration of. After 1 day of treatment, 100 μl of aluminescent cell viability assay reagent is added to each well andincubated for 10 minutes at room temperature. Luminescence is thenmeasured to determine IC₅₀.

The compound may be such that it does not modulate, or does not inhibit,TRPA1. It may not inhibit TRPA1 at an IC₅₀ of about 10 micromolar or ofabout 5 micromolar or of about 2 micromolar or of about 1 micromolar. Itmay be a TRPA1 non-inhibitor. The term not inhibit” in this context mayrefer to an inhibition of less than about 10%, or less than about 5, 2or 1% at the specified concentration.

The compounds of the present invention may inhibit phosphorylation ofco-receptor LRP6 in PA-1 teratocarcinoma cells and/or in HPAF-IIpancreatic adenocarcinoma cells by greater than about 40% after 4 hoursat a concentration of about 2 micromolar. In this context, inhibition of40% indicates that the concentration of phosphorylated LRP6 after 4hours is 40% lower than in a control to which no inhibiting compound wasadded. The inhibition under the specified conditions may be greater thanabout 40%, or greater than about 45, 50 or 55%, and may be for exampleabout 40, 45, 50, 55 or 60%. The inhibition may be achieved with aconcentration of less than about 3 micromolar, or less than about 2, 1,0.5, 0.2, 0.1 or 0.05 micromolar, or at a concentration of between about0.003 and 2 micromolar, or between about 0.003 and 1.5 micromolar, 0.003and 1 micromolar, 0.003 and 0.5 micromolar, 0.003 and 0.2 micromolar,0.003 and 0.1 micromolar, 0.003 and 0.05, 0.003 and 0.01, 0.01 and 2,0.1 and 2, 1 and 2, 0.01 and 0.1, 0.01 and 1, 0.01 and 0.1 and 0.05 or0.005 and 0.5 micromolar, e.g. at a concentration of about 0.003, 0.005,0.01, 0.002, 0.05, 0.1, 0.2, 0.5, 1, 1.5 or 2 micromolar.

The compounds of the present invention may be made as exemplified in theExamples provided herewith. A common method involves coupling1,3-dimethyl-3,7-dihydro-1H-purine-2,6-dione (or suitable derivativesuch as an acid chloride) with an amine H₂N—Ar-Cy (or a protectedderivative of that if Ar or Cy have reactive substituents other thanNH₂). This reaction may be conducted in the presence of a suitableamine, commonly a tertiary amine such as HATU and/or triethylamine.

The present invention encompasses in particular the anhydrous form ofthe free base of2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(6-phenylpyridazin-3-yl)acetamide,pharmaceutical compositions containing the anhydrous form of this freebase and methods of use of the anhydrous form of the free base in thetreatment of certain medical conditions. This compound is labelledherein as compound 5.

In the development of a drug in solid state form suitable for scale upand cGMP production and ultimately for clinical and commercial use, anacceptable level of drug activity against the target of interest is onlyone of the important variables that must be considered. For example, inthe formulation of pharmaceutical compositions it is imperative that thepharmaceutically active substance be in a form that can be reliablyreproduced in a commercial manufacturing process and which is robustenough to withstand the conditions to which the pharmaceutically activesubstance is exposed.

In a manufacturing sense it is important that during commercialmanufacture the manufacturing process of the pharmaceutically activesubstance be such that the same material is reproduced when the samemanufacturing conditions are used. In addition it is desirable that thepharmaceutically active substance exists in a solid form where minorchanges to the manufacturing conditions do not lead to major changes inthe solid form of the pharmaceutically active substance produced. Forexample it is important that the manufacturing process produce materialhaving the same crystalline properties on a reliable basis and alsoproduce material having the same level of hydration.

In addition it is important that the pharmaceutically active substancebe stable both to degradation, hygroscopicity and subsequent changes toits solid form. This is important to facilitate the incorporation of thepharmaceutically active ingredient into pharmaceutical formulations. Ifthe pharmaceutically active substance is hygroscopic (“sticky”) in thesense that it absorbs water (either slowly or over time) it is almostimpossible to reliably formulate the pharmaceutically active substanceinto a drug as the amount of substance to be added to provide the samedosage will vary greatly depending upon the degree of hydration.Furthermore variations in hydration or solid form (“polymorphism”) canlead to changes in physico-chemical properties, such as solubility ordissolution rate, which might in turn lead to inconsistent oralabsorption in a patient.

Accordingly, chemical stability, solid state stability, and “shelf life”of the pharmaceutically active agent are very important factors. In anideal situation the pharmaceutically active agent and any compositionscontaining it, should be capable of being effectively stored overappreciable periods of time, without exhibiting a significant change inthe physico-chemical characteristics of the active component such as itsactivity, moisture content, solubility characteristics, solid form andthe like.

In relation to2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(6-phenylpyridazin-3-yl)acetamide,initial studies were carried out on the free base, the preferredchemical form, and indicated that polymorphism was prevalent with thecompound being found to adopt more than one crystalline form dependingupon the manufacturing conditions. In addition it was observed that themoisture content varied from batch to batch.

Accordingly the inventors have prepared a single polymorphic form of2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(6-phenylpyridazin-3-yl)acetamidewhich overcomes or ameliorates one or more of the above identifiedproblems.

The present invention therefore encompasses an anhydrous form of thefree base (non-hydrated single polymorph) of2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-n-(6-phenylpyridazin-3-yl)acetamide.

The anhydrous free base may be crystalline. The crystalline anhydrousfree base shows on X-ray diffraction a peak on the 2theta scale at22.20°±0.5°. It also shows on X-ray diffraction peaks on the 2thetascale at 5.5°±0.5° and 14.2°±0.5°. In particular, it shows on X-raydiffraction at least four peaks on the 2theta scale selected from thegroup consisting of 5.5°±0.5° and 12.5°±0.5°, 14.20°±0.5°, 16.7°±0.5°,17.7°±0.5°, 18.8°±0.5°, 22.4°±0.5°, 24.2°±0.5° and 31.7°±0.5°.Specifically it shows on X-ray diffraction peaks on the 2theta scale ofand 5.5°±0.5° and 12.50°±0.5°, 14.2°±0.5°, 16.7°±0.5°, 17.70°±0.5°,18.0°±0.5°, 18.8°±0.5°, 19.6°±0.5°, 20.6°±0.5°, 22.4°±0.5°, 24.2°±0.5°,24.4°±0.5°, 25.0°±0.5°, 27.0°±0.5°, 27.6°±0.5°, 29.8°±0.5°, 31.7°±0.5°and 32.2°±0.5°.

The present invention also encompasses a pharmaceutical compositioncomprising the anhydrous free base as described above. It alsoencompasses a method of treating or preventing a proliferative disordercomprising administration of a therapeutically effective amount of theanhydrous free base of the invention to a patient in need thereof. Insome embodiments the proliferative disorder is cancer. It furtherencompasses the use of the anhydrous free base of the invention in thetreatment of a proliferative disorder such as cancer.

It further encompasses the use of the anhydrous free base of theinvention in the manufacture of a medicament for the treatment of aproliferative disorder. In some embodiments the proliferative disorderis cancer.

Disclosed herein are also compositions for the modulation of Wntactivity, optionally for the treatment of a disease or conditionassociated with Wnt pathway activity. These incorporate the compound ofstructure I as defined above, together with one or more pharmaceuticallyacceptable adjuvants, diluents and/or carriers.

Modulator and inhibitor compounds and agents of the present inventionmay be administered as compositions either therapeutically orpreventively. In a therapeutic application, compositions areadministered to a patient already suffering from a disease, in an amountsufficient to cure or at least partially arrest the disease and itscomplications. The composition should provide a quantity of the compoundor agent sufficient to effectively treat the patient.

The therapeutically effective dose level for any particular patient willdepend upon a variety of factors including: the disorder being treatedand the severity of the disorder; activity of the compound or agentemployed; the composition employed; the age, body weight, generalhealth, sex and diet of the patient; the time of administration; theroute of administration; the rate of sequestration of the agent orcompound; the duration of the treatment; drugs used in combination orcoincidental with the treatment, together with other related factorswell known in medicine.

One skilled in the art would be able, by routine experimentation, todetermine an effective, non-toxic amount of agent or compound whichwould be required to treat applicable diseases.

Generally, an effective dosage is expected to be in the range of about0.0001 mg to about 1000 mg per kg body weight per 24 hours; typically,about 0.001 mg to about 750 mg per kg body weight per 24 hours; about0.01 mg to about 500 mg per kg body weight per 24 hours; about 0.1 mg toabout 500 mg per kg body weight per 24 hours; about 0.1 mg to about 250mg per kg body weight per 24 hours; about 1.0 mg to about 250 mg per kgbody weight per 24 hours.

More typically, an effective dose range is expected to be in the rangeabout 1.0 mg to about 200 mg per kg body weight per 24 hours; about 1.0mg to about 100 mg per kg body weight per 24 hours; about 1.0 mg toabout 50 mg per kg body weight per 24 hours; about 1.0 mg to about 25 mgper kg body weight per 24 hours; about 5.0 mg to about 50 mg per kg bodyweight per 24 hours; about 5.0 mg to about 20 mg per kg body weight per24 hours; about 5.0 mg to about 15 mg per kg body weight per 24 hours.

Alternatively, an effective dosage may be up to about 500 mg/m².Generally, an effective dosage is expected to be in the range of about25 to about 500 mg/m², preferably about 25 to about 350 mg/m², morepreferably about 25 to about 300 mg/m², still more preferably about 25to about 250 mg/m², even more preferably about 50 to about 250 mg/m²,and still even more preferably about 75 to about 150 mg/m².

Typically, in therapeutic applications, the treatment would commonly befor the duration of the disease state.

Further, it will be apparent to one of ordinary skill in the art thatthe optimal quantity and spacing of individual dosages will bedetermined by the nature and extent of the disease state being treated,the form, route and site of administration, and the nature of theparticular individual being treated. Also, such optimum conditions canbe determined by conventional techniques.

It will also be apparent to one of ordinary skill in the art that theoptimal course of treatment, such as, the number of doses of thecomposition given per day for a defined number of days, can beascertained by those skilled in the art using conventional course oftreatment determination tests.

In general, suitable compositions may be prepared according to methodswhich are known to those of ordinary skill in the art and accordinglymay include a pharmaceutically acceptable carrier, diluent and/oradjuvant.

These compositions can be administered by standard routes. In general,the compositions may be administered by the parenteral (e.g.,intravenous, intraspinal, subcutaneous or intramuscular), oral ortopical route. More preferably administration is by the parenteralroute.

The carriers, diluents and adjuvants must be “acceptable” in terms ofbeing compatible with the other ingredients of the composition, and notdeleterious to the recipient thereof.

Examples of pharmaceutically acceptable carriers or diluents aredemineralised or distilled water; saline solution; vegetable based oilssuch as peanut oil, safflower oil, olive oil, cottonseed oil, maize oil,sesame oils, arachis oil or coconut oil; silicone oils, includingpolysiloxanes, such as methyl polysiloxane, phenyl polysiloxane andmethylphenyl polysolpoxane; volatile silicones; mineral oils such asliquid paraffin, soft paraffin or squalane; cellulose derivatives suchas methyl cellulose, ethyl cellulose, carboxymethylcellulose, sodiumcarboxymethylcellulose or hydroxypropylmethylcellulose; lower alkanols,for example ethanol or iso-propanol; lower aralkanols; lowerpolyalkylene glycols or lower alkylene glycols, for example polyethyleneglycol, polypropylene glycol, ethylene glycol, propylene glycol,1,3-butylene glycol or glycerin; fatty acid esters such as isopropylpalmitate, isopropyl myristate or ethyl oleate; polyvinylpyrridone;agar; carrageenan; gum tragacanth or gum acacia, and petroleum jelly.Typically, the carrier or carriers will form from 10% to 99.9% by weightof the compositions.

The compositions of the invention may be in a form suitable foradministration by injection, in the form of a formulation suitable fororal ingestion (such as capsules, tablets, caplets, elixirs, forexample), in an aerosol form suitable for administration by inhalation,such as by intranasal inhalation or oral inhalation, in a form suitablefor parenteral administration, that is, subcutaneous, intramuscular orintravenous injection.

For administration as an injectable solution or suspension, non-toxicparenterally acceptable diluents or carriers can include, Ringer'ssolution, isotonic saline, phosphate buffered saline, ethanol and 1,2propylene glycol.

Some examples of suitable carriers, diluents, excipients and adjuvantsfor oral use include peanut oil, liquid paraffin, sodiumcarboxymethylcellulose, methylcellulose, sodium alginate, gum acacia,gum tragacanth, dextrose, sucrose, sorbitol, mannitol, gelatine andlecithin. In addition these oral formulations may contain suitableflavouring and colourings agents. When used in capsule form the capsulesmay be coated with compounds such as glyceryl monostearate or glyceryldistearate which delay disintegration.

Adjuvants typically include emollients, emulsifiers, thickening agents,preservatives, bactericides and buffering agents.

Solid forms for oral administration may contain binders acceptable inhuman and veterinary pharmaceutical practice, sweeteners, disintegratingagents, diluents, flavourings, coating agents, preservatives, lubricantsand/or time delay agents. Suitable binders include gum acacia, gelatine,corn starch, gum tragacanth, sodium alginate, carboxymethylcellulose orpolyethylene glycol. Suitable sweeteners include sucrose, lactose,glucose, aspartame or saccharine. Suitable disintegrating agents includecorn starch, methylcellulose, polyvinylpyrrolidone, guar gum, xanthangum, bentonite, alginic acid or agar. Suitable diluents include lactose,sorbitol, mannitol, dextrose, kaolin, cellulose, calcium carbonate,calcium silicate or dicalcium phosphate. Suitable flavouring agentsinclude peppermint oil, oil of wintergreen, cherry, orange or raspberryflavouring. Suitable coating agents include polymers or copolymers ofacrylic acid and/or methacrylic acid and/or their esters, waxes, fattyalcohols, zein, shellac or gluten. Suitable preservatives include sodiumbenzoate, vitamin E, alpha-tocopherol, ascorbic acid, methyl paraben,propyl paraben or sodium bisulphite. Suitable lubricants includemagnesium stearate, stearic acid, sodium oleate, sodium chloride ortalc. Suitable time delay agents include glyceryl monostearate orglyceryl distearate.

Liquid forms for oral administration may contain, in addition to theabove agents, a liquid carrier. Suitable liquid carriers include water,oils such as olive oil, peanut oil, sesame oil, sunflower oil, saffloweroil, arachis oil, coconut oil, liquid paraffin, ethylene glycol,propylene glycol, polyethylene glycol, ethanol, propanol, isopropanol,glycerol, fatty alcohols, triglycerides or mixtures thereof.

Suspensions for oral administration may further comprise dispersingagents and/or suspending agents. Suitable suspending agents includesodium carboxymethylcellulose, methylcellulose,hydroxypropylmethyl-cellulose, poly-vinyl-pyrrolidone, sodium alginateor acetyl alcohol. Suitable dispersing agents include lecithin,polyoxyethylene esters of fatty acids such as stearic acid,polyoxyethylene sorbitol mono- or di-oleate, -stearate or -laurate,polyoxyethylene sorbitan mono- or di-oleate, -stearate or -laurate andthe like.

The emulsions for oral administration may further comprise one or moreemulsifying agents. Suitable emulsifying agents include dispersingagents as exemplified above or natural gums such as guar gum, gum acaciaor gum tragacanth.

Methods for preparing parenterally administrable compositions areapparent to those skilled in the art, and are described in more detailin, for example, Remington's Pharmaceutical Science, 15th ed., MackPublishing Company, Easton, Pa., hereby incorporated by referenceherein.

The composition may incorporate any suitable surfactant such as ananionic, cationic or non-ionic surfactant such as sorbitan esters orpolyoxyethylene derivatives thereof. Suspending agents such as naturalgums, cellulose derivatives or inorganic materials such as silicaceoussilicas, and other ingredients such as lanolin, may also be included.

The compositions may also be administered in the form of liposomes.Liposomes are generally derived from phospholipids or other lipidsubstances, and are formed by mono- or multi-lamellar hydrated liquidcrystals that are dispersed in an aqueous medium. Any non-toxic,physiologically acceptable and metabolisable lipid capable of formingliposomes can be used.

The compositions in liposome form may contain stabilisers,preservatives, excipients and the like. The preferred lipids are thephospholipids and the phosphatidyl cholines (lecithins), both naturaland synthetic. Methods to form liposomes are known in the art, and inrelation to this specific reference is made to: Prescott, Ed., Methodsin Cell Biology, Volume XIV, Academic Press, New York, N.Y. (1976), p.33 et seq., the contents of which is incorporated herein by reference.

The oral formulation may be formulated with one or morepharmacologically acceptable ingredients to make a tablet or capsuleetc. with an enteric coating. Methods for such formulations are wellknown to those skilled in the art (see e.g., Remington: The Science andPractice of Pharmacy, 19^(th) ed. (1995) Mack Publishing Company,Easton, Pa.; herein incorporated by reference). The enteric coating maybe an enteric coating which enhances delivery of the composition oractive(s) drugs to specific regions of the gastrointestinal tract forenhanced bioavailability, such as are described in United States ofAmerica Patent Application Publication No. 20040162263 entitled“Pharmaceutical formulations targeting specific regions of thegastrointesinal tract” to Sands et al and published 19 Aug. 2004.

EXAMPLES

The following examples provide compounds according to the presentinvention together with a number of general synthetic schemes forpreparing the compounds. Each synthetic scheme has been illustrated witha specific example, and the examples following that may be made by thesame general process. The person skilled in the art will readilyappreciate the variations required to the illustrated example of eachsynthetic scheme in order to prepare other related compounds.

Synthesis of Amines

Suzuki Method A: A stirred solution of the arylhalide (1 equiv.),boronic acid (1.5 equiv.) and sodium carbonate (2 equiv.) in toluene(0.08 M) and water (0.32 M) was degassed for 15 min with argon.Tetrakis(triphenylphosphine)palladium(0) (0.05 equiv.) was added toreaction mixture and the reaction mixture was heated to reflux for 16 h.After completion of starting material, the reaction mixture wasconcentrated and water was added to reaction mixture and extracted withethyl acetate. The combined organic layers were washed with brine, driedwith Na₂SO₄ and concentrated under vacuum. The crude compound waspurified by column chromatography to afford the purified product.

Synthesis of 6-phenylpyridazin-3-amine

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 7.96-7.94 (d, J=8 Hz, 2H), 7.82-7.80(d, J=9.2 Hz, 1H), 7.48-7.35 (m, 3H), 6.86-6.84 (m, 1H), 6.64 (br s,2H). LC-MS: m/z 172.0 (M+H) with a purity of 82%.

Synthesis of 4-(pyridazin-3-yl) aniline

¹H NMR (400 MHz, CDCl₃) δ (ppm): 9.01-9.0 (d, J=4.4 Hz, 1H), 8.01-7.99(d, J=8.8 Hz, 1H), 7.88-7.86 (d, J=7.9 Hz, 2H), 7.62-7.60 (m, 1H),6.68-6.66 (d, J=8.0 Hz, 2H), 5.61 (s, 2H).

Synthesis of 4-(thiazol-5-yl)aniline

¹H NMR (400 MHz, CDCl₃) δ (ppm): 8.87 (s, 1H), 8.01 (s, 1H), 7.33-7.31(d, J=8.4 Hz, 2H), 6.60-6.58 (d, J=8.3 Hz, 2H), 5.42 (s, 2H).

Synthesis of 6-(4-fluorophenyl)pyridazin-3-amine

¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 8.02-7.98 (m, 2H), 7.82 (d, J=9.2 Hz,1H), 7.29 (t, J=9.2 Hz, 2H), 6.84 (d, J=9.6 Hz, 1H), 6.5 (s, 2H). LC-MS:m/z 190 (M+H) with a purity of 99%.

Synthesis of 3′-(trifluoromethoxy)biphenyl-4-amine

¹H NMR (400 MHz, CDCl₃) δ (ppm): 7.44-7.46 (m, 1H), 7.36-7.41 (m, 4H),7.09-7.11 (m, 1H), 6.74-6.77 (m, 2H), 3.77 (bs, 2H).

Synthesis of 5-(thiazol-2-yl)pyridin-2-amine

¹H NMR (400 MHz, MeOD-d₄) δ (ppm): 8.48 (s, 1H), 7.95-7.98 (m, 1H), 7.77(d, J=3.6 Hz, 1H), 7.48 (d, J=3.6 Hz, 1H), 6.64-6.66 (m, 1H). LC-MS: m/z178 (M+H).

Synthesis of 4′-morpholinobiphenyl-4-amine

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 7.39 (d, J=8.80 Hz, 2H), 7.26 (d,J=8.80 Hz, 2H), 6.94 (d, J=8.80 Hz, 2H), 6.60 (d, J=8.80 Hz, 2H), 5.06(s, 2H), 3.74 (t, J=4.80 Hz, 4H), 3.09 (t, J=4.80 Hz, 4H).

Synthesis of 3′-morpholinobiphenyl-4-amine

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 7.32-7.34 (m, 2H), 7.19-7.23 (m, 1H),7.03 (s, 1H), 6.95-6.97 (m, 1H), 6.78-6.81 (m, 1H), 6.60-6.62 (m, 2H),3.74 (t, J=4.80 Hz, 4H), 3.14 (t, J=4.80 Hz, 4H). LC-MS: m/z 255 (M+H)with a purity of 88%.

Synthesis of 4-(pyrimidin-2-yl) aniline

Synthesis of 4-(2-chloropyrimidin-4-yl)aniline

¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 8.54 (d, J=5.2 Hz, 1H), 7.92 (d,J=8.4 Hz, 2H), 7.83 (d, J=5.2 Hz, 1H), 6.65 (d, J=8.8 Hz, 2H), 6.04 (s,2H). MS (ESI): m/z 206 [M+H]+. LC-MS: Purity of 99%.

To a stirred solution of 4-(2-chloropyrimidin-4-yl)aniline (1 equiv.) inmethanol (0.1 M) and 10% aqueous NaOH (0.24 M) was added 10% Pd/C (20wt. %) and stirred under Hydrogen balloon pressure at room temperaturefor 16 h. The reaction mixture was filtered through celite pad andwashed with methanol, filtrate concentrated under reduced pressure. Theresultant solid was recrystallized with 30% ethyl acetate in petroleumether to afford 4-(pyrimidin-4-yl)aniline as a pale yellow solid. ¹H-NMR(400 MHz; DMSO-d₆) δ (ppm): 9.02 (s, 1H), 8.62 (d, J=5.2 Hz, 1H), 7.94(d, J=8.4 Hz, 2H), 7.81 (d, J=5.6 Hz, 1H), 6.65 (d, J=8.8 Hz, 2H), 5.79(s, 2H). MS (ESI): m/z 172 [M+H]+. LC-MS: Purity of 94%.

Suzuki Method B: A solution of aryl halide (1 equiv.) in 1,4-dioxane(0.12 M) and water (0.5 M) was treated with the respective boronic acidor ester (1.2 equiv.), tricyclohexylphosphine (0.1 equiv.) and K₃PO₄ (2equiv.) at room temperature. Nitrogen gas was passed through thereaction mixture for 15 min. Pd₂(dba)₃ (0.1 equiv.) was added to thereaction mixture and degassed for another 15 min. The reaction mixtureheated to 100° C. for 16 h. After completion, reaction mixture wascooled to room temperature, added water, extracted with ethyl acetatethrice. The combined organic layers were washed with brine solution,dried over anhydrous Na₂SO₄, filtered, rotary evaporated and dried undervacuum to afford crude product. The crude product was purified by columnchromatography to afford the purified product.

Synthesis of 2,3′-bipyridin-6′-amine

MS (ESI): m/z 172.13 [M+H]+.

Synthesis of 4-(thiazol-2-yl) aniline

¹H-NMR (400 MHz; CDCl₃) δ (ppm): 7.77 (d, J=6.0 Hz, 1H), 7.19 (d, J=3.6Hz, 1H), 6.71 (d, J=4.8 Hz, 2H), 3.8 (brs, 2H). LC-MS: m/z 175 [M−H]−.Purity of 62%.

Suzuki Method C: A stirred solution of arylhalide (1.1 equiv.) in1,4-dioxane (0.7 M) and water (3.5 M) was treated with the respectiveboronic acid or ester (1 equiv.) and K₂CO₃ (2.4 equiv.) at roomtemperature. Nitrogen gas was passed through the reaction mixture for 15min. Pd(dppf)Cl₂.DCM (0.02 equiv.) was added to the reaction mixture anddegassed for another 15 minutes. The reaction mixture heated to 100° C.for 4 h. After completion, reaction mixture was cooled to roomtemperature, added water, extracted with ethyl acetate trice. Thecombined organic layers was washed with brine solution, dried overanhydrous Na₂SO₄, filtered, rotary evaporated and dried under vacuum toafford crude product. The crude product was purified by columnchromatography to afford the purified product.

Synthesis of 4-(thiophen-3-yl) aniline

¹H-NMR (400 MHz; CDCl₃) δ (ppm): 7.41 (d, J=8.4 Hz, 2H), 7.33 (d, J=3.1Hz, 1H), 7.32 (d, J=0.9 Hz, 1H), 7.31 (s, 1H), 6.71 (d, J=8.4 Hz, 2H),3.7 (brs, 2H). MS (ESI): m/z 176 [M+H]+. LC-MS: Purity of 97%.

Synthesis of 6-(pyridin-4-yl) pyridazin-3-amine

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 7.96-7.94 (d, J=8 Hz, 2H), 7.82-7.80(d, J=9.2 Hz, 1H), 7.48-7.35 (m, 3H), 6.86-6.84 (m, 1H), 6.64 (br s,2H). LC-MS: m/z 172.0 (M+H) with a purity of 82%.

Synthesis of 5-(2-methylthiazol-4-yl)pyridin-2-amine

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 8.58-8.57 (1H, d, J=1.6 Hz),7.94-7.92 (1H, dd, J1=2 Hz, J2=6.4 Hz), 7.15 (1H, s), 6.56-6.54 (1H, d,J=8.4 Hz), 4.54 (2H, brs), 2.76 (3H, s).

Synthesis of N-methyl-4-(thiophen-3-yl)aniline

¹H NMR (400 MHz, CDCl₃) δ (ppm): 7.45-7.43 (d, J=7.0 Hz, 2H), 7.33 (s,1H), 7.32-7.27 (m, 3H), 6.65-6.63 (d, J=7.1 Hz, 1H), 3.77-3.58 (brs,1H), 2.87 (s, 1H).

Synthesis of 4-(oxazol-2-yl) aniline

To a solution of commercially available 2-(4-nitrophenyl)oxazole, 1 (1equiv.) in methanol (0.05 M) was added Pd/C (10% by wt) and stirredreaction at room temperature under H2 gas balloon pressure for 4 h.After completion of starting material, the reaction mixture was filteredthrough Celite bed and filtrate was concentrated to give 4-(oxazol-2-yl)aniline, 2 as a brown solid. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 8.0 (s,1H), 7.64 (d, J=8.4 Hz, 2H), 7.20 (s, 1H), 6.63-6.61 (d, J=8.4 Hz, 2H),5.68 (brs, 2H). LC-MS: m/z 161.0 (M+H) with a purity of 97%.

Synthesis of 5-phenylthiophen-2-amine

Step 1: Preparation of ethyl 2-amino-5-phenylthiophene-3-carboxylate.

A stirred solution of the respective aldehyde or ketone (1 equiv.),ethyl cyano acetate (1 equiv.) and S powder in ethanol (1.6 M) wastreated with morpholine (5 M) dropwise at room temperature and stirredfor 3 h. After completion of starting material, the reaction mixture wasconcentrated and crude compound was purified by column chromatography togive the product.

¹H NMR (400 MHz, CDCl₃) δ (ppm): 7.45-7.43 (m, 2H), 7.34-7.30 (m, 2H),7.27-7.20 (m, 2H), 6.0 (brs, 2H), 4.33-4.27 (q, J=7.2 Hz, 2H), 1.39-1.35(t, J=7.2 Hz, 3H). LC-MS: m/z 248.2 (M+H) with a purity of 94%.

Step 2: Preparation of 5-phenylthiophen-2-amine

A solution of ethyl 2-amino-5-phenylthiophene-3-carboxylate (1 equiv.)in ethanol (0.04 M) was added 50% aq. HCl (0.04 M) and reaction mixturewas heated to reflux for 4 h. After completion of starting material, thereaction mixture was cooled to room temperature and concentrated undervacuum and basified with aq.NaHCO₃ solution and extracted with ethylacetate twice. The combined organic layers were washed with brinesolution dried over NaSO₄ concentrated under vacuum. The crude compoundwas purified by column chromatography to give the purified product.LC-MS: m/z 176.8 (M+H) with a purity of 61%.

Synthesis of 4-phenylthiophen-2-amine

Step 1: Preparation of ethyl 2-amino-4-phenylthiophene-3-carboxylate

¹H NMR (400 MHz, CDCl₃) δ (ppm): 7.30-7.28 (m, 5H), 6.07 (brs, 1H), 6.05(brs, 2H), 4.06-4.00 (q, J=6.8 Hz, 2H), 0.94-0.91 (t, J=6.8 Hz, 3H).LC-MS: m/z 248.03 (M+H) with a purity of 98%.

Step 2: Preparation of 4-phenylthiophen-2-amine

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 7.60-7.58 (m, 2H), 7.40-7.38 (m, 3H),7.29-7.27 (m, 2H), 7.15-7.11 (m, 2H). LC-MS: m/z 176.0 (M+H) with apurity of 94%.

Synthesis of 4-(pyrrolidin-1-yl)aniline

Step 1: Preparation of 1-(4-nitrophenyl)pyrrolidine

Potassium carbonate (2 equiv.) and the respective amine (1.1 equiv.)were added to a stirred solution of 1-fluoro-4-nitrobenzene, 1 (1equiv.) in anhydrous DMSO (0.5 M) and stirred at 120° C. for 18 h. Uponconsumption of starting material, the reaction was diluted with waterand extracted with dichloromethane. The combined organic layers werewashed with brine, dried over Na₂SO₄ and concentrated under vacuum.Crude product was purified using column chromatography to afford thepurified product. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 8.05 (d, J=9.2 Hz,2H), 6.62 (d, J=9.2 Hz, 2H), 3.38 (m, 4H), 1.99 (m, 4H). LC-MS: m/z 193(M+H)

Step 2: Preparation of 4-(pyrrolidin-1-yl)aniline

1-(4-nitrophenyl)pyrrolidine was dissolved in ethyl acetate (0.05 M) andreduced with H-cube at 50° C., 10 bar. Reduction was completed in 2cycles. The solvent was evaporated off in vacuo, and the crude purifiedusing-column chromatography to afford the product. ¹H NMR (400 MHz,DMSO-d₆) δ (ppm): 6.49 (d, J=8.4 Hz, 2H), 6.34 (d, J=8.4 Hz), 4.24 (s,2H), 3.07 (m, 4H), 1.87 (m, 4H). LC-MS: m/z 163 (M+H).

Synthesis of (S)-tert-butyl4-(4-aminophenyl)-3-methylpiperazine-1-carboxylate

Step 1: Preparation of (S)-tert-butyl3-methyl-4-(4-nitrophenyl)piperazine-1-carboxylate

¹H NMR (400 MHz, CDCl₃) δ (ppm): 8.15-8.12 (m, 2H), 6.79-6.77 (d, J=9.6Hz, 2H), 4.13-3.96 (m, 3H), 3.57-3.53 (m, 1H), 3.29-3.22 (m, 2H), 3.12(br s, 1H), 1.49 (s, 9H), 1.18 (d, J=6.4 Hz, 3H). LC-MS: m/z 363(M+H+41).

Step 2: Preparation of (S)-tert-butyl4-(4-aminophenyl)-3-methylpiperazine-1-carboxylate

¹H NMR (400 MHz, CDCl₃) δ (ppm): 6.84 (d, J=8.6 Hz, 2H), 6.64 (d, J=8.6Hz, 2H), 3.53 (br s, 3H), 3.26 (br s, 2H), 2.99-2.96 (m, 1H), 2.88-2.85(m, 1H), 1.48 (s, 9H), 0.87 (d, J=6.4 Hz, 3H). LC-MS: m/z 292 (M+H).

Synthesis of (R)-tert-butyl4-(4-aminophenyl)-3-methylpiperazine-1-carboxylate

Step 1: Preparation of (R)-tert-butyl3-methyl-4-(4-nitrophenyl)piperazine-1-carboxylate

¹H NMR (400 MHz, CDCl₃) δ (ppm): 8.15-8.13 (m, 2H), 6.79-6.77 (m, 2H),4.12 (br s, 2H), 4.02 (br s, 1H), 3.57-3.53 (m, 1H), 3.29-3.22 (m, 2H),3.12 (br s, 1H), 1.49 (s, 9H), 1.18 (d, J=6.8 Hz, 3H). LC-MS: m/z 363(M+H+41).

Step 2: Preparation of (R)-tert-butyl4-(4-aminophenyl)-3-methylpiperazine-1-carboxylate

¹H NMR (400 MHz, CDCl₃) δ (ppm): 6.84 (d, J=8.4 Hz, 2H), 6.65-6.63 (m,2H), 3.53 (br s, 3H), 3.26 (br s, 2H), 2.99-2.96 (m, 1H), 2.88-2.85 (m,1H), 1.48 (s, 9H), 0.87 (d, J=6 Hz, 3H). LC-MS: m/z 292 (M+H).

Amide coupling Method A: To a stirred solution of commercially available2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetic acid, 1(1 equiv.) in dichloromethane (0.01 M) was added HATU (1.3 equiv.),triethylamine (1.5 equiv.) and the respective amine (1 equiv.). Thereaction mixture was allowed to stir at room temperature. Uponcompletion of the reaction, water was added to the reaction mixture andthe mixture was extracted with dichloromethane. The organic layer waswashed with brine solution, dried over anhydrous Na₂SO₄, andconcentrated under vacuum to afford the crude product. The crude productis further purified by column chromatography.

Compound 1:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiazol-5-yl)phenyl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.58 (s, 1H), 9.03 (s, 1H), 8.24 (s,1H), 8.08 (s, 1H), 7.65 (s, 4H), 5.20 (s, 2H), 3.40 (s, 3H), 3.20 (s,3H).

Compound 2:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-phenylthiophen-2-yl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 11.71 (s, 1H), 8.09 (s, 1H),7.55-7.53 (d, J=8 Hz, 2H), 7.38-7.34 (t, J=7.6 Hz, 2H), 7.28-7.21 (m,2H), 6.73-6.72 (d, J=3.6 Hz, 1H), 5.25 (s, 2H), 3.46 (s, 3H), 3.20 (s,3H). LC-MS: m/z 396.03 (M+H) with a purity of 99.02%. HPLC: At 254 nmwith a purity of 95%.

Compound 3:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-phenylthiophen-2-yl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 11.70 (s, 1H), 8.09 (s, 1H),7.64-7.62 (d, J=7.6 Hz, 2H), 7.42-7.38 (t, J=7.2 Hz, 2H), 7.31-7.28 (m,2H), 7.08-7.07 (d, J=1.6 Hz, 1H), 5.26 (s, 2H), 3.46 (s, 3H), 3.20 (s,3H). LC-MS: m/z 396.11 (M+H) with a purity of 99%.

Compound 4:N-(4-(1H-imidazol-1-yl)phenyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide

¹H NMR (400 MHz, DMSO-d₆) 3 (ppm): 10.57 (s, 1H), 8.18 (s, 1H), 8.08 (s,1H), 7.68-7.70 (m, 3H), 7.59-7.61 (m, 2H), 7.08 (s, 1H), 5.22 (s, 2H),3.46 (s, 3H), 3.20 (s, 3H). LC-MS: m/z 380 (M+H) with a purity of 99%.

Compound 5:2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(6-phenylpyridazin-3-yl)acetamide(alternatively named2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(6-phenylpyridazin-3-yl)acetamide)

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 11.77 (s, 1H), 8.32-8.25 (m, 2H),8.12-8.10 (m, 3H), 7.57-7.49 (m, 3H), 5.37 (s, 2H), 3.46 (s, 3H), 3.19(s, 3H). LC-MS: m/z 390.1 (M+H) with a purity of 99%.

Compound 6: 2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(pyridazin-3-yl)phenyl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.66 (s, 1H), 9.17-9.16 (d, J=4.4,1H), 8.20-8.14 (m, 3H), 8.09 (s, 1H), 7.77-7.73 (m, 3H), 5.25 (s, 2H),3.47 (s, 3H), 3.27 (s, 3H), 3.20 (s, 3H). LC-MS: m/z at 392 [M+H] with99%.

Compound 7:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(pyridazin-3-yl)phenyl)acetamide

¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 10.65 (brs, 1H), 9.18 (s, 1H), 8.80(d, J=5.2 Hz, 1H), 8.20 (d, J=8.8 Hz, 2H), 8.07 (s, 1H), 8.02 (d, J=5.2Hz, 1H), 7.75 (d, J=8.8 Hz, 2H), 5.25 (s, 2H), 3.46 (s, 3H), 3.20 (s,3H). MS (ESI): m/z 392 [M+H]+. LC-MS: Purity of 97%.

Compound 8: 2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(pyrimidin-2-yl)phenyl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.66 (s, 1H), 8.87-8.86 (d, J=4.0Hz, 2H), 8.37-8.35 (d, J=8.4 Hz, 2H), 8.08 (s, 1H,), 7.74-7.72 (d, J=8.8Hz, 2H), 7.40-7.38 (t, J=4.8 Hz, 1H), 5.24 (s, 2H), 3.46 (s, 3H), 3.20(s, 3H). LC-MS: m/z at 392 [M+H] with 99%.

Compound 9:N-(2,3′-bipyridin-6′-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide

¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 11.21 (s, 1H), 9.08 (s, 1H), 8.67 (d,J=3.4 Hz, 1H), 8.47 (d, J=7.2 Hz, 1H), 8.08 (s, 1H), 8.03 (d, J=7.2 Hz,2H), 7.9 (t, J=7.6 Hz, 1H), 7.38 (t, J=5.2 Hz, 1H), 5.30 (s, 2H), 3.46(s, 3H), 3.19 (s, 3H). MS (ESI): m/z 392.13 [M+H]+. LC-MS: Purity of97%.

Compound 10:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(pyridin-4-yl)phenyl)acetamide

¹H NMR (400 MHz, MeOD-d₄) δ (ppm): 8.55 (d, J=6.00 Hz, 2H), 7.992 (s,1H), 7.73-7.78 (m, 4H), 7.71 (d, J=6.00 Hz, 2H), 5.28 (s, 2H), 3.58 (s,3H), 3.33 (s, 3H). LC-MS: m/z 391 (M+H) with a purity of 99%.

Compound 11:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(6-(4-fluorophenyl)pyridazin-3-yl) acetamide

¹H-NMR (400 MHz; CDCl₃) δ (ppm): 11.65 (s, 1H), 8.58 (d, J=9.6 Hz, 1H),7.99 (dd, J=5.2 Hz, J=3.6 Hz, 2H), 7.90 (d, J=9.2 Hz, 1H), 7.73 (s, 1H),7.17 (t, J=8.8 Hz, 1H), 5.57 (s, 2H), 3.64 (s, 3H), 3.42 (s, 3H). MS(ESI): m/z 410.17 [M+H]+. LC-MS: Purity of 99%.

Compound 12:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(6-(pyridin-4-yl)pyridazin-3-yl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 11.88 (brs, 1H), 8.76 (s, 2H),8.39-8.37 (m, 2H), 8.15-8.09 (m, 3H), 5.38 (s, 2H), 3.46 (s, 3H), 3.19(s, 3H). LC-MS: m/z 393.20 (M+H) with a purity of 98.93%. HPLC: At 254nm with a purity of 98%.

Compound 13:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(3-methylbiphenyl-4-yl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 9.79 (s, 1H), 8.10 (s, 1H), 7.63 (d,J=7.6 Hz, 2H), 7.54-7.40 (m, 5H), 7.34 (t, J=7.2 Hz, 1H), 5.26 (s, 2H),3.45 (s, 3H), 3.22 (s, 3H), 2.32 (s, 3H).

Compound 14: 2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2-methoxybiphenyl-4-yl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.56 (s, 1H), 8.09 (s, 1H),7.48-7.30 (m, 5H), 7.28-7.17 (m, 3H), 5.23 (s, 2H), 3.72 (s, 3H), 3.46(s, 3H), 3.20 (s, 3H).

Compound 15: (S)-tert-butyl4-(4-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamido)phenyl)-3-methylpiperazine-1-carboxylate

¹H NMR (400 MHz, CDCl₃) δ (ppm): 9.27 (s, 1H), 7.77 (s, 1H), 7.42 (d,J=8.8 Hz, 2H), 6.85 (d, J=8.8 Hz, 2H), 4.94 (s, 2H), 4.00-3.78 (br s,1H), 3.71-3.69 (m, 2H), 3.61 (s, 3H), 3.46 (s, 3H), 3.42-3.36 (m, 1H),3.21 (br s, 1H), 3.05-3.03 (m, 2H), 1.47 (s, 9H), 0.94 (d, J=6.4 Hz,3H). LC-MS: m/z 512 (M+H) with a purity of 97%.

Compound 16: (R)-tert-butyl4-(4-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamido)phenyl)-3-methylpiperazine-1-carboxylate

¹H NMR (400 MHz, CDCl₃) δ (ppm): 9.35 (br s, 1H), 7.77 (s, 1H),7.46-7.44 (m, 2H), 6.90 (br s, 2H), 4.94 (s, 2H), 3.89 (br s, 1H),3.70-3.68 (m, 1H), 3.67-3.61 (m, 4H), 3.46 (m, 4H), 3.32-3.25 (br s,1H), 3.09 (br s, 2H), 1.48 (s, 9H), 0.96 (d, J=6.4 Hz, 3H). LC-MS: m/z512 (M+H) with a purity of 97%.

Compound 17:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-methyl-N-(4-(thiophen-3-yl)phenyl)acetamide

¹H NMR (400 MHz, CDCl₃) δ (ppm): 7.74-7.72 (d, J=7.6 Hz, 2H), 7.54-7.51(d, J=9.2 Hz, 2H), 7.43-7.26 (m, 4H), 4.89 (s, 2H), 3.59 (s, 3H), 3.38(s, 3H), 3.34 (s, 3H). MS (ESI) m/z 410 [M+1].

Amide coupling Method B: To a stirred solution of commercially available2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetic acid, 1in N,N-dimethylformamide (0.2 M) was added Hunig's base (1.5 equiv.),HATU (1.5 equiv.) and the respective amine (1.2 equiv.). The mixture wasstirred at room temperature. After consumption of starting material, thereaction mixture was quenched with water and extracted withdichloromethane. The combined organic layers were washed with brinesolution, dried over Na₂SO₄ and concentrated under vacuum to afford thecrude product. The crude product is further purified by columnchromatography.

Compound 18:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.43 (s, 1H), 8.07 (s, 1H),7.78-7.77 (m, 1H), 7.69-7.67 (m, 2H), 7.62-7.60 (m, 3H), 7.53-7.52 (m,1H), 5.22 (s, 2H), 3.47 (s, 3H), 3.21 (s, 3H). LC-MS: m/z 396 (M+H) witha purity of 98%.

Compound 19:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-phenylpyridin-2-yl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 11.07 (bs, 1H), 8.68 (s, 1H),8.04-8.12 (m, 3H), 7.70-7.72 (m, 2H), 7.48 (t, J=7.60 Hz, 2H), 7.38 (t,J=7.60 Hz, 1H), 5.29 (s, 2H), 3.19 (s, 3H). LC-MS: m/z 391 (M+H) with apurity of 98%.

Compound 20: tert-butyl4-(4-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanamido)phenyl)piperazine-1-carboxylate

¹H NMR (600 MHz, C₆D₆) δ (ppm): 9.84 (s, 1H), 8.39 (d, J=9.0 Hz, 1H),7.92 (d, J=2.9 Hz, 1H), 7.01 (s, 1H), 6.64 (dd, J=9.0, 2.9 Hz, 1H), 4.35(s, 2H), 3.26 (m, 4H), 3.26 (s, 3H), 3.25 (s, 3H), 2.53-2.40 (m, 4H),1.45 (s, 10H). LC-MS: m/z 499 (M+H) with a purity of 99%

Compound 21:N-(5-(4-(3-chlorobenzyl)piperazin-1-yl)pyridin-2-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide

¹H NMR (600 MHz, CDCl₃) δ (ppm): 9.43 (s, 1H), 8.01-7.94 (m, 2H), 7.74(s, 1H), 7.36 (s, 1H), 7.24 (ddd, J=12.1, 7.6, 3.3 Hz, 4H), 5.09 (s,2H), 3.60 (s, 3H), 3.54 (s, 2H), 3.42 (s, 3H), 3.22-3.14 (m, 4H),2.65-2.57 (m, 4H). LC-MS: m/z 523 (M+H), 521(M−H) with a purity of 98%.

Compound 22: tert-butyl4-(4-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamido)phenyl)piperazine-1-carboxylate

¹H NMR (600 MHz, CDCl₃) δ (ppm): 9.30 (s, 1H), 7.77 (s, 1H), 7.42 (d,J=9.1 Hz, 2H), 6.87 (d, J=8.9 Hz, 2H), 4.95 (s, 2H), 3.60 (s, 3H), 3.57(m, 4H), 3.45 (s, 3H), 3.07 (m, 4H), 1.47 (s, 9H). LC-MS: m/z 498 (M+1),496 (M−1) with purity of 99%.

Compound 23: tert-butyl4-(4-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanamido)phenyl)piperazine-1-carboxylate

¹H NMR (600 MHz, MeOD-d₄) δ (ppm): 8.23 (s, 1H), 7.83 (s, 2H), 7.51 (d,J=7.6 Hz, 2H), 7.00 (d, J=7.6 Hz, 2H), 5.81 (q, J=7.2 Hz, 1H), 3.64 (bs,7H), 3.39 (s, 3H), 3.15 (m, 4H), 1.96 (d, J=7.2 Hz, 3H), 1.55 (s, 9H).LC-MS: m/z 512 (M+1), 510(M−1) with purity of 98%.

Amide coupling Method C: To a stirred solution of commercially available2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetic acid, 1in dichloromethane (0.1 M) was added the respective amine (100 1equiv.), EDCI (1.2 equiv.) and HOBT (1.2 equiv.). The reaction mixturewas stirred at room temperature for 16 h. After completion of startingmaterial, water was added to the reaction mixture and product wasextracted with 10% methanol/chloroform twice. The organic layer wasdried over anhydrous Na2SO4, concentrated under vacuum to afford thecrude product. The crude product is further purified by columnchromatography.

Compound 24:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(oxazol-2-yl)phenyl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.72 (s, 1H), 8.19 (s, 1H), 8.09 (s,1H), 7.96-7.94 (d, J=8.8 Hz, 2H), 7.75-7.72 (d, J=8.8 Hz, 2H), 7.35 (s,1H), 5.24 (s, 2H), 3.46 (s, 3H), 3.19 (s, 3H). LC-MS: m/z 379.3 (M+H)with a purity of 96.48%. HPLC: At 254 nm with a purity of 97%.

Amide coupling Method D: A stirred solution of commercially available2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetic acid, 1(1 equiv.) in N,N-dimethylforamide (0.18 M), was added Hunig's base (1.5equiv.), HOBt (1.5 equiv.), EDCI (1.5 equiv.) and the respective amine(1.5 equiv.). The mixture was stirred at room temperature for 16 h.After consumption of starting material, the reaction mixture wasquenched with water and extracted with dichloromethane. The combinedorganic layers were washed with brine solution, dried over Na₂SO₄ andconcentrated under vacuum. The crude compound was purified by columnchromatography to afford the product.

Compound 25:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(pyridin-2-yl)phenyl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.57 (s, 1H), 8.61-8.62 (m, 1H),8.05-8.08 (m, 3H), 7.91 (d, J=8.00 Hz, 1H), 7.82-7.86 (m, 1H), 7.68 (d,J=8.00 Hz, 1H), 7.28-7.31 (m, 1H), 5.23 (s, 2H), 3.46 (s, 3H), 3.20 (s,3H).

Compound 26:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(pyridin-3-yl)phenyl)acetamide

¹H NMR (400 MHz, DMSO-d₆) (ppm): 10.56 (s, 1H), 8.87 (s, 1H), 8.52-8.53(m, 1H), 8.08 (s, 1H), 8.04-8.06 (m, 1H), 7.68-7.73 (m, 4H), 7.44-7.47(m, 1H), 5.23 (s, 2H), 3.46 (s, 3H), 3.20 (s, 3H). LC-MS: m/z 391 (M+H)with a purity of 97%.

Compound 27:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2′-methoxybiphenyl-4-yl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.48 (s, 1H), 8.08 (s, 1H),7.60-7.58 (m, 2H), 7.44-7.42 (m, 2H), 7.33-7.25 (m, 2H), 7.08 (d, J=8Hz, 1H), 7.00 (t, J=7.4 Hz, 1H), 5.21 (s, 2H), 3.74 (s, 3H), 3.20 (s,3H). LC-MS: m/z 420 (M+H) with a purity of 98%

Compound 28:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2′-methylbiphenyl-4-yl)acetamide

¹H NMR (400 MHz, MeOD-d₄) δ (ppm): 8.00 (s, 1H), 7.63-7.61 (m, 2H),7.28-7.23 (m, 3H), 7.22-7.19 (m, 2H), 7.18-7.15 (m, 1H), 5.28 (s, 2H),3.58 (s, 3H), 3.35 (s, 3H), 2.24 (s, 3H). LC-MS: m/z 404 (M+H) with apurity of 98%.

Compound 29:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(3′-methylbiphenyl-4-yl)acetamide

¹H NMR (400 MHz, MeOD-d₄) δ (ppm): 7.99 (s, 1H), 7.65-7.63 (m, 2H),7.58-7.56 (m, 2H), 7.41 (s, 1H), 7.38-7.36 (m, 1H), 7.29 (t, J=7.8, 1H),7.14-7.12 (m, 1H), 5.27 (s, 2H), 3.58 (s, 3H), 3.34 (s, 3H), 2.39 (s,3H). LC-MS: m/z 404 (M+H) with a purity of 97%.

Compound 30:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4′-methylbiphenyl-4-yl)acetamide

¹H NMR (400 MHz, MeOD-d₄) δ (ppm): 7.99 (s, 1H), 7.64-7.62 (m, 2H),7.57-7.55 (m, 2H), 7.48 (d, J=8 Hz, 2H), 7.23 (d, J=8 Hz, 2H), 5.27 (s,2H), 3.58 (s, 3H), 3.34 (s, 3H), 2.36 (s, 3H). LC-MS: m/z 404 (M+H) witha purity of 97%.

Compound 31:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4′-(trifluoromethoxy)biphenyl-4-yl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.54 (s, 1H), 8.08 (s, 1H), 7.76 (d,J=8.40 Hz, 2H), 7.65-7.70 (m, 4H), 7.42 (d, J=8.40 Hz, 2H), 5.23 (s,2H), 3.46 (s, 3H), 3.20 (s, 3H). LC-MS: m/z 474 (M+H) with a purity of98%.

Compound 32:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(3′-methoxybiphenyl-4-yl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.52 (s, 1H), 8.08 (s, 1H), 7.65 (s,4H), 7.35 (t, J=8 Hz, 1H), 7.21 (d, J=8 Hz, 1H), 7.16 (m, 1H), 6.91-6.88(m, 1H), 5.22 (s, 2H), 3.81 (s, 3H), 3.20 (s, 3H). LC-MS: m/z 420 (M+H)with a purity of 99%.

Compound 33:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(2-methylthiazol-4-yl)phenyl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.51 (s, 1H), 8.08 (s, 1H), 7.89 (d,J=8.8 Hz, 2H), 7.81 (s, 1H), 7.63 (d, J=8.8 Hz, 2H), 5.22 (s, 2H), 3.46(s, 3H), 3.20 (s, 3H), 2.70 (s, 3H). LC-MS: m/z 411.5 (M+H) with apurity of 96%.

Compound 34:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiazol-2-yl)phenyl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.67 (s; 1H), 8.08 (s, 1H),7.94-7.91 (m, 2H), 7.88 (d, J=3.2 Hz, 1H), 7.73-7.72 (m, 2H), 7.70 (m,1H), 5.24 (s, 2H), 3.47 (s, 3H), 3.20 (s, 3H). LC-MS: m/z 397 (M+H) witha purity of 95%.

Amide coupling Method E: A solution of commercially available2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetic acid, 1(1.3 equiv.) and triethylamine (2.1 equiv.) in dichloromethane (0.1 M)was cooled to 0° C. under nitrogen atmosphere and treated with isobutylchloroformate (2.0 equiv.). The reaction mixture stirred for 30 minutes,treated with the respective amine (1.0 equiv.) and gently brought up toroom temperature for 2-18 h until judged complete by LC-MS. It waspartitioned between dichloromethane and saturated sodium bicarbonate.The organic phase was separated, washed with aqueous sodium chloride,dried over sodium sulfate and concentrated to dryness. The residue waspurified using preparative HPLC to give the product.

Compound 35:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(6-phenylpyridin-3-yl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.74 (s, 1H), 8.81-8.82 (m, 1H),8.09-8.12 (m, 2H), 8.03-8.05 (m, 2H), 7.94-7.96 (m, 1H), 7.45-7.49 (m,2H), 7.39-7.41 (m, 1H), 5.26 (s, 2H), 3.46 (s, 3H), 3.20 (s, 3H). LC-MS:m/z 391 (M+H) with a purity of 96%.

Compound 36:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-phenylpyrazin-2-yl)acetamide

¹H NMR (400 MHz, MeOD-d₄) δ (ppm): 9.35 (bs, 1H), 8.86 (d, J=1.2 Hz,1H), 8.00-8.02 (m, 3H), 7.42-7.52 (m, 3H), 5.37 (s, 2H), 3.58 (s, 3H),3.33 (s, 3H). LC-MS: m/z 392 (M+H) with a purity of 99%.

Compound 37:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(4-fluorophenyl)pyridin-2-yl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 11.08 (bs, 1H), 8.67 (s, 1H),8.06-8.11 (m, 3H), 7.75-7.78 (m, 2H), 7.29-7.33 (m, 2H), 5.29 (s, 2H),3.46 (s, 3H), 3.19 (s, 3H). LC-MS: m/z 409 (M+H) with a purity of 98%.

Compound 38: methyl4′-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamido)biphenyl-4-carboxylate

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.57 (s, 1H), 8.08 (s, 1H), 8.01 (d,J=8.40 Hz, 2H), 7.81 (d, J=8.40 Hz, 2H), 7.69-7.76 (m, 4H), 5.23 (s,211), 3.86 (s, 3H), 3.46 (s, 3H), 3.20 (s, 3H). LC-MS: m/z 448 (M+H)with a purity of 97%.

Compound 39:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(3-phenylisoxazol-5-yl) acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 12.28 (s, 1H), 8.08 (s, 1H), 7.82 (s,2H), 7.49 (s, 3H), 6.68 (s, 1H), 5.29 (s, 2H), 3.46 (s, 3H), 3.19 (s,3H). LC-MS: m/z 381 (M+H) with a purity of 97%.

Compound 40:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(oxazol-5-yl)phenyl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.59 (s, 1H), 8.39 (s, 1H), 8.07 (s,1H), 7.68 (m, 4H), 7.59 (s, 1H), 5.22 (s, 2H), 3.46 (s, 3H), 3.20 (s,3H). LC-MS: m/z 381 (M+H) with a purity of 95%.

Compound41:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(thiazol-2-yl)pyridin-2-yl)acetamide

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 11.27 (s, 1H), 8.93 (d, J=2 Hz, 1H),8.32 (dd, J=8.8, 2.4 Hz, 1H), 8.11-8.09 (m, 1H), 8.07 (s, 1H), 7.95 (d,J=3.2 Hz, 1H), 7.82 (d, J=3.2 Hz, 1H), 5.31 (s, 2H), 3.46 (s, 3H), 3.19(s, 3H). LC-MS: m/z 398 (M+H) with a purity of 96%.

Removal of Boc protecting group: To a mixture of tert-butyl4-(6-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamido)pyridin-3-yl)piperazine-1-carboxylate, 1 (1 equiv.) indichloromethane (0.1 M) was added trifluroacetic acid (10 equiv.) atroom temperature. The reaction mixture was allowed to stir for 2 hfollowed by which the volatiles were evaporated. The resulting mass wasthen subjected to purification on a preparative TLC.

Compound 42:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(piperazin-1-yl)pyridin-2-yl)acetamide

¹H NMR (600 MHz, CDCl₃: MeOD (10:1)) δ (ppm): 7.86 (d, J=8.9 Hz, 1H),7.83 (d, J=2.8 Hz, 1H), 7.69 (s, 1H), 7.20-7.16 (m, 1H), 5.06 (s, 2H),3.47 (s, 3H), 3.25 (s, 3H), 3.22 (dt, J=3.9, 1.6 Hz, 1H), 3.14 (dd,J=6.2, 3.9 Hz, 3H), 3.07-3.03 (m, 4H). LC-MS: m/z 399 (M+H) with apurity of 96%.

Compound 43:(S)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(2-methylpiperazin-1-yl)phenyl)acetamide

¹H NMR (400 MHz, CDCl₃) δ (ppm): 9.37 (s, 1H), 7.77 (s, 1H), 7.45 (d,J=8.8 Hz, 2H), 6.92 (d, J=8.8 Hz, 2H), 4.95 (s, 2H), 3.61 (m, 4H), 3.47(s, 3H), 3.24-3.06 (m, 5H), 2.91-2.87 (m, 1H), 0.99 (d, J=6.4 Hz, 3H).LC-MS: m/z 412 (M+H) with a purity of 99%.

Compound 44:(R)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-2-methylpiperazin-1-yl)phenyl)acetamide

¹H NMR (400 MHz, CDCl₃) δ (ppm): 9.37 (s, 1H), 7.77 (s, 1H), 7.45 (d,J=8.8 Hz, 2H), 6.92 (d, J=8.8 Hz, 2H), 4.95 (s, 2H), 3.61 (m, 4H), 3.47(s, 3H), 3.24-3.06 (m, 5H), 2.91-2.87 (m, 1H), 0.99 (d, J=6.4 Hz, 3H).LC-MS: m/z 412 (M+H) with a purity of 99%.

Acylation Method A: To a solution of2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(piperazin-1-yl)pyridin-2-yl)acetamide,1 (1 equiv.) in dry N,N-dimethylformamide (0.07 M) was added Hunig'sbase (2 equiv.) and the respective acid chloride (1 equiv.). Thereaction was stirred for 16 h at room temperature then quenched by theaddition of distilled water. The solvent was removed in vacuo, and thecrude material was partitioned between chloroform and saturated NaHCO₃.The crude product was further purified by column chromatography.

Compound 45:N-(5-(4-benzoylpiperazin-1-yl)pyridin-2-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide

1H NMR (600 MHz, CDCl₃) δ (ppm): 9.53 (s, 1H), 8.02 (d, J=8.9 Hz, 1H),7.98 (d, J=1.9 Hz, 1H), 7.73 (s, 1H), 7.43 (s, 5H), 7.27 (dd, J=9.1, 3.0Hz, 1H), 5.11 (s, 2H), 3.93 (br s, 2H), 3.60 (m, 5H), 3.41 (s, 3H), 3.16(br d, J=59.6 Hz, 4H). LC-MS: 503 (M+H), 524 (M+Na) with a purity of92%.

Compound 46:(S)—N-(4-(4-benzoyl-2-methylpiperazin-1-yl)phenyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide

¹H NMR (400 MHz, CDCl₃) δ (ppm): 9.31 (s, 1H), 7.76 (s, 1H), 7.44-7.42(m, 7H), 6.87 (d, J=8.8 Hz, 2H), 4.94 (s, 2H), 3.61 (m, 4H), 3.46 (m,5H), 1.55 (s, 7H). LC-MS: m/z 516 (M+H) with a purity of 98%.

Compound 47:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(4-(3,3-dimethylbutanoyl)piperazin-1-yl)pyridin-2-yl)acetamide

¹H NMR (600 MHz, C₆D₆) δ (ppm): δ 9.97 (s, 1H), 8.44 (d, J=9.1 Hz, 1H),7.94 (d, J=2.9 Hz, 1H), 7.09 (s, 1H), 6.71 (dd, J=9.1, 3.0 Hz, 1H), 4.49(s, 2H), 3.55 (s, 2H), 3.29 (s, 3H), 3.27 (s, 3H), 2.89 (s, 2H), 2.45(d, J=39.6 Hz, 4H), 2.03 (s, 2H), 1.10 (s, 9H). LC-MS: m/z 497 (M+H),495 (M−H) with a purity of 97%.

Compound 48:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(piperazin-1-yl)pyridin-2-yl)acetamide

¹H NMR (600 MHz, C₆D₆) δ (ppm): δ 9.50 (s, 1H), 8.47 (d, J=9.2 Hz, 1H),7.88 (d, J=2.6 Hz, 1H), 6.86 (s, 1H), 6.66 (dd, J=9.1, 3.0 Hz, 1H), 4.12(s, 2H), 3.47-3.42 (m, 2H), 3.23 (s, 6H), 3.01 (s, 3H), 2.66-2.59 (m,2H), 2.39-2.34 (m, 2H), 2.25-2.21 (m, 2H). LC-MS: m/z 439 (M−H), 441(M+H) with a purity of 97%.

Compound 49:N-(5-(4-(4-chlorobenzoyl)piperazin-1-yl)pyridin-2-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide

¹H NMR (600 MHz, CDCl₃) δ (ppm): 9.53 (s, 1H), 8.03 (d, J=9.0 Hz, 1H),7.97 (d, J=2.9 Hz, 1H), 7.73 (s, 1H), 7.40 (m, J=8.7 Hz, 4H), 7.28-7.26(m, 1H), 5.11 (s, 2H), 3.91 (s, 2H), 3.60 (s, 4H), 3.48 (s, 1H), 3.41(s, 3H), 3.17 (bs, 4H). LC-MS: m/z 537 (M+1) and 535 (M−1) with a purityof 98%.

Compound 50:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(4-(thiophene-2-carbonyl)piperazin-1-yl)pyridin-2-yl)acetamide

¹H NMR (600 MHz, DMSO-d₆) δ (ppm): 10.77 (s, 1H), 8.07 (d, J=3.0 Hz,1H), 8.06 (s, 1H), 7.86 (d, J=9.0 Hz, 1H), 7.78 (dd, J=5.0, 1.1 Hz, 1H),7.46 (dd, J=3.6, 1.0 Hz, 1H), 7.44 (dd, J=9.1, 2.9 Hz, 1H), 7.15 (dd,J=5.0, 3.6 Hz, 1H), 5.22 (s, 2H), 3.79 (m, 4H), 3.45 (s, 3H), 3.22 (m,4H), 3.19 (s, 3H). LC-MS: m/z 509 (M+H), 507 (M−H) with a purity of 99%.

Acylation Method B: To a mixture of the2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(piperazin-1-yl)pyridin-2-yl)acetamide,1 (1 equiv.) and the respective acid (1 equiv.) in N,N-dimethylforamide(0.06 M), HATU (1.2 equiv.) and Hunig's base (3 equiv.) in a dropwisefashion at room temperature. The reaction mixture was allowed to stirfor 30 min followed by which it was quenched with saturated NaHCO₃solution. The resulting mixture was extracted with ethyl acetate twiceand the combined organic layer was evaporated and the crude productswere subjected to purification on a preparative TLC plate.

Compound 51:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(4-(3-fluorobenzoyl)piperazin-1-yl)pyridin-2-yl)acetamide

¹H NMR (600 MHz, DMSO-d₆) δ (ppm): 10.80 (s, 1H), 8.09-8.02 (m, 2H),7.85 (d, J=9.1 Hz, 1H), 7.52 (td, J=7.9, 5.8 Hz, 1H), 7.43 (dd, J=9.1,3.0 Hz, 1H), 7.36-7.22 (m, 3H), 5.21 (s, 2H), 3.76 (bs, 2H), 3.45 (s,3H), 3.24 (bs, 2H), 3.18 (s, 3H), 3.13 (bs, 2H). LC-MS: m/z 521 (M+1),519 (M−1) with a purity of 95%.

Compound 52:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(4-(4-fluorobenzoyl)piperazin-1-yl)pyridin-2-yl)acetamide

¹H NMR (600 MHz, CDCl₃) δ (ppm): 9.69 (s, 1H), 8.04 (d, J=9.1 Hz, 1H),7.96 (d, J=2.8 Hz, 1H), 7.74 (s, 1H), 7.50-7.40 (m, 2H), 7.30 (dd,J=9.2, 3.0 Hz, 1H), 7.15-7.08 (m, 2H), 5.12 (s, 2H), 3.90 (bs, 3H), 3.66(bs, 1H), 3.60 (s, 3H), 3.41 (s, 3H), 3.17 (s, 4H). LC-MS: m/z 519 (M−H)and purity of 98%.

Alkylation Procedure: To a mixture of the2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(piperazin-1-yl)pyridin-2-yl)acetamide,1 (1 equiv.) in N,N-dimethylforamide (0.08 M) and Hunig's base was addedthe respective arylbromide (1 equiv.) at room temperature. The reactionmixture was allowed to stir for 24 h followed by evaporation of thevolatiles and addition water. The resulting mixture was extracted withethyl acetate trice and the organic layer was evaporated and the crudeproducts were subjected to purification on a preparative TLC plate.

Compound 53:N-(5-(4-benzylpiperazin-1-yl)pyridin-2-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide

¹H NMR (600 MHz, CDCl₃) δ (ppm): 9.64 (s, 1H), 8.01 (d, J=2.9 Hz, 1H),7.99 (d, J=9.1 Hz, 1H), 7.76 (s, 1H), 7.37 (m, 5H), 7.33 (d, J=4.3 Hz,1H), 5.10 (s, 2H), 3.59 (s, 3H), 3.57 (s, 2H), 3.40 (s, 3H), 3.22-3.13(m, 4H), 2.71-2.45 (m, 4H). LC-MS: 489 (M+H) with a purity of 94%.

Compound 54:N-(5-(4-(3,5-difluorobenzyl)piperazin-1-yl)pyridin-2-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide

¹H NMR (600 MHz, CDCl₃) δ (ppm): 9.33 (s, 1H), 8.02-7.94 (m, 2H), 7.73(d, J=13.6 Hz, 1H), 7.25-7.22 (m, 1H), 6.90 (t, J=13.6 Hz, 2H),6.74-6.65 (m, 1H), 5.13-5.04 (m, 2H), 3.63-3.59 (m, 3H), 3.57-3.52 (m,2H), 3.45-3.39 (m, 3H), 3.18 (dd, J=14.6, 9.6 Hz, 4H), 2.62 (s, 4H).LC-MS: m/z 525 (M+H), 523 (M−H) with purity of 97%.

Reductive amination procedure: To a mixture of the2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(piperazin-1-yl)pyridin-2-yl)acetamide,1 (1 equiv.) and the respective aldehyde (1 equiv.) in dichloroethane(0.1 M) was added acetic acid (2 equiv.) followed by triacetoxy sodiumborohydride (1.4 equiv.) at room temperature. The reaction was stirredfor an hour and then quenched with saturated NaHCO₃ (1 mL). The organicswere extracted in ethyl acetate trice and the combined organic layerswere dried. The residue was purified by using preparative TLC.

Compound 55:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(4-(2-fluorobenzyl)piperazin-1-yl)pyridin-2-yl)acetamide

¹H NMR (600 MHz, CDCl₃) δ (ppm): 9.61 (s, 1H), 7.99-7.98 (m, 1H), 7.96(d, J=9.2 Hz, 1H), 7.73 (s, 1H), 7.45-7.38 (m, 1H), 7.29-7.24 (m, 1H),7.22 (dd, J=9.1, 3.0 Hz, 1H), 7.15-7.10 (m, 1H), 7.07-7.02 (m, 1H), 5.10(s, 2H), 3.67 (s, 2H), 3.59 (s, 3H), 3.40 (s, 3H), 3.24-3.13 (m, 4H),2.67 (s, 4H). LC-MS: m/z 507 (M+1), 505(M−1) with purity of 96%.

Compound 56:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(4-(4-fluorobenzyl)piperazin-1-yl)pyridin-2-yl)acetamide

¹H NMR (600 MHz, CDCl₃) δ (ppm): 9.29 (s, 1H), 7.98 (d, J=9.1 Hz, 1H),7.95 (d, J=2.4 Hz, 1H) 7.73 (s, 1H), 7.38 (m, 2H), 7.26-7.20 (m, 1H),7.04 (t, J=8.6 Hz, 2H), 5.08 (s, 2H), 3.64 (bs, 2H), 3.60 (s, 3H), 3.42(s, 3H), 3.25 (bs, 4H), 2.70 (bs, 4H). LC-MS: m/z 507 (M+1), 505(M−1)with a purity of 96%.

Compound 57:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(4-(3-fluorobenzyl)piperazin-1-yl)pyridin-2-yl)acetamide

¹H NMR (600 MHz, CDCl₃) δ (ppm): 9.48 (s, 1H), 8.01 (s, 1H), 7.99 (m,1H), 7.76 (s, 1H), 7.34-7.29 (m, 1H), 7.28-7.23 (m, 1H), 7.15-7.09 (m,2H), 6.99 (ddd, J=7.8, 5.2, 1.8 Hz, 1H), 5.12 (s, 2H), 3.65-3.61 (m,3H), 3.62-3.58 (m, 2H), 3.44 (s, 3H), 3.25-3.16 (m, 4H), 2.66 (m, 4H).LC-MS: m/z 507 (M+H), 505 (M−H) with a purity of 95%.

Mitsunobu Reaction Conditions:

To a stirred solution of commercially available1,3-dimethyl-1H-purine-2,6(3H,7H)-dione, 1 (2 equiv.), (S)-ethyl2-hydroxypropanoate, 2 (1 equiv.) and triphenyl phosphine (1.45 g,5.555) in tetrahydrofuran (0.14 M) was added DIAD (2 equiv.) dropwise atroom temperature, and the resulting reaction mixture was stirred for 5h. After completion of starting material, the reaction mixture wasdiluted with water and extracted with ethyl acetate twice. The combinedorganic layers were washed with brine, dried over anhydrous Na₂SO₄, andconcentrated under vacuum. The crude compound was purified by columnchromatography to afford the product.

Alkylation Method A:

To a solution of commercially available1,3-dimethyl-1H-purine-2,6(3H,7H)-dione, 1 (1 equiv.) inN,N-dimethylformamide (0.28 M) was added methyl2-bromo-2-methylpropanoate, 2 (1.2 equiv.), K₂CO₃ (2 equiv.) at roomtemperature and warmed to 80° C. for 20 h. After completion, thereaction mixture was cooled to room temperature; water was added andmixture was extracted with ethyl acetate trice. The combined ethylacetate layers were washed with water, brine, dried over anhydrousNa₂SO₄, filtered, rotary evaporated and dried under vacuum to afford theproduct.

Alkylation Method B:

A solution of 1,3-dimethyl-1H-purine-2,6(3H,7H)-dione, 2 (1 equiv.) and2-bromo-N-(4-(thiophen-3-yl)phenyl)butanamide, 1 (1.1 equiv.) inN,N-dimethylformamide (0.1 M) cooled to 0° C. under nitrogen atmosphere,was treated with sodium hydride (2.5 equiv., 60% mineral oil).

The mixture was stirred for 18 h at room temperature under nitrogenatmosphere and was partitioned between dichloromethane and water. Theorganic phase was separated, washed with aqueous sodium chloride, driedover sodium sulfate and concentrated to dryness. The residue waspurified using preparative HPLC to give the final product.

Hydrolysis Conditions:

To a stirred solution of (R)-ethyl2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanoate, 3 (1equiv.) in tetrahydrofuran (0.18 M), methanol (0.36 M) and water (0.36M) was added LiOH.H₂O (1.5 equiv.) at room temperature and resultingreaction mixture was stirred at room temperature for 2 h. Aftercompletion of starting material, the reaction mixture was concentratedand the residue was dissolved in water and washed with ethyl acetatetwice and acidified with aq.KHSO₄; product was extracted with 10%methanol/chloroform twice. The combined organic layers were dried usingNa₂SO₄, concentrated under vacuum to give the product.

Compound 58(R)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)propanamide

Step 1: Preparation of (R)-ethyl2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanoate

Intermediate was prepared using the Mitsunobu conditions as described.¹H NMR (400 MHz, CDCl₃) δ (ppm): 7.75 (s, 1H), 5.63-5.58 (q, J=6.8 Hz,1H), 4.26-4.24 (q, J=6.8 Hz, 2H), 3.61 (s, 3H), 3.39 (s, 3H), 1.86-1.84(d, J=6.8 Hz, 3H), 1.29-1.25 (t, J=6.8 Hz, 3H). LC-MS: m/z 281.3 (M+H)with a purity of 43%.

Step 2: Preparation of(R)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl) propanoicacid

Intermediate was prepared using the hydrolysis condition as described.¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 13.25 (brs, 1H), 8.20 (s, 1H),5.50-5.44 (q, J=7.6 Hz, 1H), 3.44 (s, 3H), 3.24 (s, 3H), 1.76-1.74 (d,J=7.6 Hz, 3H). LC-MS: m/z 250.9 (M+H) with a purity of 97%.

Step 3: Preparation of(R)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)propanamide

The final product was prepared using amide coupling method A reactionconditions as described. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.47 (s,1H), 8.33 (s, 1H), 7.79 (s, 1H), 7.69-7.67 (d, J=8.8 Hz, 2H), 7.62-7.60(d, J=8.4 Hz, 3H), 7.53-7.52 (d, J=4.8 Hz, 1H), 5.71-5.69 (q, J=6.9 Hz,1H), 3.46 (s, 3H), 3.20 (s, 3H), 1.84-1.83 (d, J=6.8 Hz, 3H). LC-MS: m/z410.13 (M+H) with a purity of 99%.

Compound 59:(S)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)propanamide

Step 1: Preparation of (S)-ethyl2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanoate

Intermediate was prepared using the Mitsunobu conditions as described.¹H NMR (400 MHz, CDCl₃) δ (ppm): 7.75 (s, 1H), 5.63-5.58 (q, J=6.8 Hz,1H), 4.26-4.24 (q, J=6.8 Hz, 2H), 3.61 (s, 3H), 3.39 (s, 3H), 1.86-1.84(d, J=6.8 Hz, 3H), 1.29-1.25 (t, J=6.8 Hz, 3H). LC-MS: m/z 281.4 (M+H)with a purity of 32%.

Step 2: Preparation of(S)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl) propanoicacid

Intermediate was prepared using the hydrolysis condition as described.¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 13.25 (brs, 1H), 8.20 (s, 1H),5.50-5.44 (q, J=7.6 Hz, 1H), 3.44 (s, 3H), 3.24 (s, 3H), 1.76-1.74 (d,J=7.6 Hz, 3H). LC-MS: m/z 226.14 (M+H) with a purity of 95%.

Step 3:(S)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)propanamide

The final product was prepared using amide coupling method A reactionconditions as described. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.47 (s,1H), 8.33 (s, 1H), 7.79 (s, 1H), 7.69-7.67 (d, J=8.8 Hz, 2H), 7.62-7.60(d, J=8.4 Hz, 3H), 7.53-7.52 (d, J=4.8 Hz, 1H), 5.71-5.69 (q, J=6.9 Hz,1H), 3.46 (s, 3H), 3.20 (s, 3H), 1.84-1.83 (d, J=6.8 Hz, 2H). LC-MS: m/z410.07 (M+H) with a purity of 99%. ee: 99.67%.

Compound 60:(S)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)propanamide

Step 1: Preparation of methyl2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-2-methylpropanoate

Intermediate was prepared using alkylation method A conditions asdescribed. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 8.24 (s, 1H), 3.62 (s,3H), 3.44 (s, 3H), 3.19 (s, 3H), 1.80 (s, 6H). MS (ESI): m/z 281 [M+H]+.LC-MS: Purity of 34%.

Step 2: Preparation of2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-2-methylpropanoic acid

Intermediate was prepared using hydrolysis conditions as described.¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 13.10 (brs, 1H), 8.22 (s, 1H), 3.45(s, 3H), 3.21 (s, 3H), 1.81 (s, 6H). MS (ESI): m/z 267 [M+H]+. LC-MS:Purity of 87%.

Step 3: Preparation of2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-2-methyl-N-(4-(thiophen-3-yl)phenyl)propanamide

The final product was prepared using amide coupling method A reactionconditions as described. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 9.37 (s,1H), 8.28 (s, 1H), 7.78 (s, 1H), 7.65-7.60 (m, 3H), 7.60-7.52 (m, 3H),3.47 (s, 3H), 3.17 (s, 3H), 1.90 (s, 6H). MS (ESI): m/z 424.05 [M+H]+.LC-MS: Purity of 96%.

Compound 61:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)propanamide

Step 1: Preparation ethyl2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl) propanoate

Intermediate was prepared using alkylation method A conditions asdescribed. ¹H NMR (400 MHz, CDCl₃) δ (ppm): 7.97 (s, 1H), 5.59 (q,J=7.60 Hz, 1H), 4.24 (q, J=7.20 Hz, 2H), 3.58 (s, 3H), 3.36 (s, 3H),1.83 (d, J=7.60 Hz, 3H), 1.28 (t, J=7.20 Hz, 3H). LC-MS: m/z 281 (M+H).

Step 2: Preparation2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanoic acid

Intermediate was prepared using hydrolysis conditions as described. ¹HNMR (400 MHz, DMSO-d₆) δ (ppm): 8.19 (s, 1H), 5.46 (q, J=7.20 Hz, 1H),3.44 (s, 3H), 3.21 (s, 3H), 1.74 (d, J=7.20 Hz, 3H). LC-MS: m/z 253(M+H).

Step 3: Preparation2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)propanamide

The final product was prepared using amide coupling method D reactionconditions as described. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.46 (s,1H), 8.31 (s, 1H), 7.78 (d, J=1.60 Hz, 1H), 7.60-7.68 (m, 5H), 7.52 (d,J=4.80 Hz, 1H), 5.69 (q, J=7.20 Hz, 1H), 3.45 (s, 3H), 3.20 (s, 3H),1.83 (d, J=7.20 Hz, 1H). LC-MS: m/z 410 (M+H) with a purity of 97%.

Compound 62:N-(biphenyl-4-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanamide

The final product was prepared using amide coupling method D reactionconditions as described. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.49 (s,1H), 8.32 (s, 1H), 7.62-7.68 (m, 6H), 7.43 (t, J=7.60 Hz, 2H), 7.32 (t,J=7.60 Hz, 1H), 5.70 (q, J=7.20 Hz, 1H), 3.46 (s, 3H), 3.20 (s, 3H),1.84 (d, J=7.20 Hz, 3H). LC-MS: m/z 404 (M+H) with a purity of 99%.

Compound 63:N-(4-(4-(3,5-difluorobenzyl)piperazin-1-yl)phenyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanamide

Product was prepared using alkylation method A reaction conditions asdescribed. ¹H NMR (600 MHz, CDCl₃) δ (ppm): 9.25 (s, 1H), 7.86 (s, 1H),7.42 (d, J=9.1 Hz, 2H), 6.92 (d, J=6.3 Hz, 2H), 6.85 (d, J=9.1 Hz, 2H),6.70 (m, 1H), 5.55 (q, J=7.1 Hz, 1H), 3.60 (s, 3H), 3.56 (s, 2H), 3.46(s, 3H), 3.17 (m, 4H), 2.62 (m, 4H), 1.87 (d, J=7.1 Hz, 3H). LC-MS: m/z538 (M+1) with HPLC purity of 99%.

Compound 64:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)butanamide

Step 1: Preparation 2-bromo-N-(4-(thiophen-3-yl)phenyl)butanamide

Intermediate was prepared using amide coupling method E reactionconditions as described. ¹H NMR (400 MHz, CDCl₃) δ (ppm): 7.58-7.59 (m,4H), 7.42 (s, 1H), 7.36-7.40 (m, 2H), 4.43-4.48 (m, 1H), 2.22-2.31 (m,1H), 2.07-2.20 (m, 1H), 1.10-1.14 (m, 3H).

Step 2: Preparation2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)butanamide

Final product was prepared using the alkylation method B reactionconditions as described. ¹H NMR (400 MHz, MeOD-d₄) δ (ppm): 8.26 (s,1H), 7.61 (s, 4H), 7.56-7.57 (m, 1H), 7.42-7.46 (m, 2H), 5.67-5.71 (m,1H), 3.56 (s, 3H), 3.36 (s, 3H), 2.23-2.38 (m, 2H), 1.04-1.07 (m, 3H).LC-MS: m/z 424 (M+H) with a purity of 97%.

Compound 65:N-(biphenyl-4-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-3-methylbutanamide

Step 1: Preparation N-(biphenyl-4-yl)-2-bromo-3-methylbutanamide

Intermediate was prepared using amide coupling method B reactionconditions as described. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.40 (s,1H), 7.63-7.70 (m, 6H), 7.42-7.46 (m, 2H), 7.31-7.35 (m, 1H), 4.30-4.33(m, 1H), 2.19-2.28 (m, 1H), 1.11 (d, J=6.80 Hz, 3H), 0.99 (d, J=6.80 Hz,3H). LC-MS: m/z 404 (M+H).

Step 2: PreparationN-(biphenyl-4-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-3-methylbutanamide

The product was prepared using alkylation method A reaction conditionsas described. ¹H NMR (400 MHz, MeOD-d₄) δ (ppm): 8.36 (s, 1H), 7.66-7.69(m, 2H), 7.58-7.60 (m, 4H), 7.41 (t, J=7.60 Hz, 2H), 7.30 (t, J=7.60 Hz,1H), 5.52-5.54 (m, 1H), 3.56 (s, 3H), 3.38 (s, 3H), 2.57-2.67 (m, 1H),1.16 (d, J=6.80 Hz, 3H), 0.96 (d, J=6.80 Hz, 3H). LC-MS: m/z 432 (M+H)with a purity of 99%.

Preparation of 1,3,8-trimethyl-1H-purine-2,6(3H, 7H)-dione

Step 1: Preparation ofN-(5-amino-1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropyrimidin-4-yl)acetamide

To a stirred solution of commercially available5,6-diamino-1,3-dimethylpyrimidine-2,4(1H,3H)-dione, 1 (1 equiv.) inacetic acid (4 equiv.) at room temperature and warmed to 70° C. for 4 h.The reaction mixture was cooled to room temperature then diluted withice water and concentrated under reduced pressure to get crude compound,dried under vacuum. The resultant crude was precipitated with 20%dichloromethane in hexane to affordN-(5-amino-1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropyrimidin-4-yl)acetamideas a yellow solid. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 8.36 (s, 1H), 6.58(s, 2H), 3.30 (s, 3H), 3.10 (s, 3H), 1.92 (s, 3H). MS (ESI): m/z 213[M+H]+. LC-MS: Purity of 96%.

Step 2: Preparation of 1,3,8-trimethyl-1H-purine-2,6(3H,7H)-dione

To a stirred solid ofN-(5-amino-1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropyrimidin-4-yl)acetamide,2 at room temperature and warmed to 250° C. for 2 h. The reactionmixture was cooled to room temperature and the reaction mixture wasprecipitated with 30% dichloromethane in hexane to afford1,3,8-trimethyl-1H-purine-2,6(3H,7H)-dione as a yellow solid. ¹H-NMR(400 MHz; DMSO-d₆) δ (ppm): 12.50 (brs, 1H), 3.40 (s, 3H), 3.22 (s, 3H),2.37 (s, 3H). MS (ESI): m/z 195 [M+H]+. LC-MS: Purity of 96%.

Compound 66:N-(4-(thiophen-3-yl)phenyl)-2-(1,3,8-trimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide

Step 1: Preparation of ethyl2-(1,3,8-trimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetate

Intermediate was prepared using alkylation method A reaction conditionsas described. ¹H-NMR (400 MHz; CDCl₃) δ (ppm): 5.09 (s, 2H), 4.27 (q,J=7.2 Hz), 3.58 (s; 3H), 3.38 (s, 3H), 2.43 (s, 3H), 1.31 (t, J=7.2 Hz,3H). MS (ESI): m/z 281 [M+H]+. LC-MS: Purity of 93%.

Step 2: Preparation of2-(1,3,8-trimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetic acid

Intermediate was prepared using hydrolysis reaction conditions asdescribed. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 13.34 (brs, 1H), 5.08 (s,2H), 3.41 (s, 3H), 3.20 (s, 3H), 2.38 (s, 3H). MS (ESI): m/z 251 [M−H]−.LC-MS: Purity of 94%.

Step 3: Preparation ofN-(4-(thiophen-3-yl)phenyl)-2-(1,3,8-trimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide

Product was prepared using amide coupling method A reaction conditionsas described. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 10.50 (s, 1H), 7.80 (s,1H), 7.70-7.61 (m, 5H), 7.53 (d, J=4.4 Hz, 1H), 5.22 (s, 2H), 3.44 (s,3H), 3.20 (s, 3H), 2.42 (s, 3H). MS (ESI): m/z 408.5 [M−H]−. LC-MS:Purity of 96%.

Compound 67: (S)—N-(4-(thiazol-2-yl)phenyl)-2-(1,3,8-trimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanamide

Step 1: Preparation of (S)-ethyl2-(1,3,8-trimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanoate

Intermediate was prepared using Mitsunobu reaction conditions asdescribed. ¹H-NMR (400 MHz; CDCl₃) δ (ppm): 5.50 (q, J=7.2 Hz, 1H), 4.11(q, J=3.6 Hz, 2H), 3.41 (s, 3H), 3.19 (s, 3H), 2.46 (s, 3H), 1.66 (d,J=7.2 Hz, 3H), 1.14 (t, J=7.2 Hz, 3H). LC-MS: m/z 295.1 (M+H) with apurity of 57.79% (desired). LCMS: m/z 279 (M+H) with a purity of 31%.

Step 2: Preparation of(S)-2-(1,3,8-trimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanoic acid

Intermediate was prepared using hydrolysis reaction conditions asdescribed. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 13.03 (brs, 1H), 5.43 (q,J=6.8 Hz, 1H), 3.41 (s, 3H), 3.20 (s, 3H), 2.45 (s, 3H), 1.66 (d, J=7.2Hz, 3H). LC-MS: m/z 267.1 [M+H]+. LC-MS: Purity of 98%.

Step 3: Preparation of (S)—N-(4-(thiazol-2-yl)phenyl)-2-(1,3,8-trimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanamide

Product was prepared using amide coupling method A reaction conditionsas described. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 9.99 (s, 1H), 7.89-7.82(m, 3H), 7.71-7.65 (m, 3H), 5.74 (q, J=, 6.9 Hz, 1H), 3.46 (s, 3H), 3.33(s, 3H), 3.19 (s, 3H), 1.77 (d, J=7.2 Hz, 3H). LC-MS: m/z 422.9[M−H]−(95%) and 90% ee.

Preparation of ethyl2-(8-bromo-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl) acetate

To a solution of commercially available8-bromo-1,3-dimethyl-1H-purine-2,6(3H,7H)-dione (1 equiv.) inN,N-dimethylformamide (0.2 M) was added bromoethylacetate (1.2 equiv.),K₂CO₃ (2.5 equiv.) at room temperature and warmed to 70° C. for 4 h. Thereaction mixture was cooled to room temperature then poured into icewater to precipitate the crude compound. The solid was filtered andwashed with water, dried under vacuum. The resultant solid wasre-crystallized with isopropanol to afford the product. ¹H-NMR (400 MHz;CDCl₃) δ (ppm): 5.12 (s, 2H), 4.28 (q, J=7.6 Hz, 2H), 3.58 (s, 3H), 3.38(s, 3H), 1.32 (t, J=6.8 Hz, 3H). MS (ESI): m/z 331 [M+H]+.

Compound 68:N-(4-(thiophen-3-yl)phenyl)-2-(1,3,8-trimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide

Step 1: Preparation of2-(8-cyclopropyl-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetic acid

Intermediate was prepared using Suzuki Method C reaction conditions asdescribed. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 13.30 (brs, 1H), 5.20 (s,2H), 3.37 (s, 3H), 3.19 (s, 3H), 2.15-2.12 (m, 1H), 1.05 (d, J=2.8 Hz,2H), 0.98 (d, J=2.8 Hz, 2H). MS (ESI): m/z 279 [M+H]+. LC-MS: Purity of88%.

Step 2: Preparation of2-(8-cyclopropyl-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Product was prepared using amide coupling reaction conditions asdescribed. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 10.46 (s, 1H), 7.78 (s,1H), 7.69-7.52 (m, 5H), 7.52 (d, J=4.4 Hz, 1H), 5.32 (s, 2H), 3.39 (s,3H), 3.19 (s, 3H), 2.15 (m, 1H), 1.06-1.02 (m, 4H). MS (ESI): m/z 436[M+H]+. LC-MS: Purity of 94%.

Compound 69:N-(4-(thiophen-3-yl)phenyl)-2-(1,3,8-trimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide

Step 1: Preparation of2-(1,3-dimethyl-2,6-dioxo-8-phenyl-2,3-dihydro-1H-purin-7(6H)-yl) aceticacid

Intermediate was prepared using Suzuki Method C reaction conditions asdescribed. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 13.38 (brs, 1H), 7.67-7.57(m, 5H), 5.01 (s, 2H), 3.49 (s, 3H), 3.25 (s, 3H). MS (ESI): m/z 315[M+H]+. LC-MS: Purity of 94%.

Step 2: Preparation of2-(1,3-dimethyl-2,6-dioxo-8-phenyl-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Product was prepared using amide coupling reaction conditions asdescribed. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 10.53 (s, 1H), 7.80 (s,1H), 7.72-7.67 (m, 5H), 7.61-7.53 (m, 6H), 5.21 (s, 2H), 3.51 (s, 3H),3.24 (s, 3H). MS (ESI): m/z 472 [M+H]+. LC-MS: Purity of 98%.

Compound 70:2-(1,3-dimethyl-2,6-dioxo-8-(trifluoromethyl)-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Step 1: Preparation ofN-(5-amino-1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropyrimidin-4-yl)-2,2,2-trifluoroacetamide

To a stirred solution of commercially available5,6-diamino-1,3-dimethylpyrimidine-2,4(1H,3H)-dione (1 equiv.) inbenzene (0.06 M) at room temperature was added trifluoroacetic acid (1equiv.) and the resulting reaction mixture was heated to reflux for 4 h.After completion of the starting material, the reaction mixture wascooled to room temperature and concentrated under vacuum. The residuewas washed with diethyl ether and dried to give the intermediateN-(5-amino-1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropyrimidin-4-yl)-2,2,-trifluoroacetamide as a brown solid. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm):9.76 (s, 1H), 6.97 (s, 2H), 3.32 (s, 3H), 3.11 (s, 3H). LC-MS: m/z 264.9(M−H) with a purity of 84%.

Step 2: Preparation of1,3-dimethyl-8-(trifluoromethyl)-1H-purine-2,6(3H, 7H)-dione

N-(5-amino-1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropyrimidin-4-yl)-2,2,2-trifluoroacetamidewas mixed with P₂O₅(w/w) and heated to 200° C. for 15 h. The black masswas cooled to room temperature and quenched with ice and extracted withethyl acetate twice. The combined ethyl acetate layers were dried overNa₂SO₄ and concentrated under vacuum. The crude compound was purified bycolumn chromatography to afford the intermediate1,3-dimethyl-8-(trifluoromethyl)-1H-purine-2,6(3H,7H)-dione as a lightbrown solid. ¹H NMR (400 MHz, DMSO-d₆) (ppm): 3.34 (s, 3H), 3.26 (brs,1H), 3.24 (s, 3H). LC-MS: m/z 249.0 (M+H) with a purity of 88%.

Step 3: Preparation of ethyl2-(1,3-dimethyl-2,6-dioxo-8-(trifluoromethyl)-2,3-dihydro-1H-purin-7(6H)-yl)acetate

Intermediate was prepared using alkylation method A reaction conditionsas described. ¹H NMR (400 MHz, CDCl₃) δ (ppm): 5.28 (s, 2H), 4.31-4.25(q, J=7.2 Hz, 2H), 3.61 (s, 3H), 3.40 (s, 3H), 1.29-1.21 (t, J=7.2 Hz,3H).

Step 4: Preparation of2-(1,3-dimethyl-2,6-dioxo-8-(trifluoromethyl)-2,3-dihydro-1H-purin-7(6H)-yl)aceticacid

Intermediate was prepared using hydrolysis reaction conditions asdescribed. LC-MS: m/z 307.3 (M+H) with a purity of 41%.

Step 5: Preparation of2-(1,3-dimethyl-2,6-dioxo-8-(trifluoromethyl)-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Product was prepared using amide coupling method A reaction conditionsas described. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.57 (s, 1H),7.79-7.70 (m, 1H), 7.68-7.66 (d, J=8.4 Hz, 2H), 7.61-7.55 (m, 3H),7.52-7.50 (dd, J1=3.2 Hz, J2=1.6 Hz, 1H), 5.43 (s, 2H), 3.45 (s, 3H),3.22 (s, 3H). LC-MS: m/z 464.02 (M+H) with a purity of 97%.

Compound 71:2-(2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Step 1: Preparation of 2-(2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)aceticacid

Commercially available 1H-purine-2,6(3H,7H)-dione (1 equiv.) in water(1.5 M) at room temperature was added 2M NaOH solution (0.65 M) and theresulting solution was stirred for 30 min and chloroacetic acid (1equiv.) was added and resulting reaction mixture was refluxed for 5 h.The reaction mixture was cooled to room temperature and stirred for 16h. The precipitated solid was removed by filtration and water wasacidified with conc HCl (pH 2). The solid collected by filtration,washed with hot ethanol to afford the intermediate2-(2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetic acid as a white solid.¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 13.35 (brs, 1H), 11.59 (s, 1H), 10.88(brs, 1H), 7.91 (s, 1H), 5.0 (s, 2H). LC-MS: m/z 211.1 (M+H) with apurity of 97%.

Step 2: Preparation of2-(2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Product was prepared using amide coupling method D reaction conditionsas described. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 11.60 (s, 1H), 10.88(brs, 1H), 10.43 (brs, 1H), 7.95 (s, 1H), 7.80-7.79 (d, J=2 Hz, 1H),7.70-7.67 (d, J=8.8 Hz, 2H), 7.61-7.59 (m, 3H), 7.53-7.52 (d, J=4.8 Hz,1H), 5.14 (s, 2H). LC-MS: m/z 366.10 (M−H) with a purity of 95.43%.HPLC: At 278 nm with a purity of 96%.

Compound 72:2-(1,3-diethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Step 3: Preparation of2-(1,3-diethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

A stirred solution of2-(2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide(1 equiv.)) in N,N-dimethylformamide (0.03 M) was added K₂CO₃ (2.5equiv.) and stirred at room temperature for 15 min. Ethyl iodide (2.5equiv.) was added to the reaction mixture and stirred for another 4 h atroom temperature. After completion, water was added to the reactionmixture and extracted with ethyl acetate twice. The combined organiclayer was washed with brine, dried over anhydrous Na₂SO₄, andconcentrated under vacuum and purified by column chromatography toafford the product2-(1,3-diethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamideas an off-white solid. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.48 (s, 1H),8.08 (s, 1H), 7.70-7.68 (d, J=8.4 Hz, 2H), 7.62-7.61 (m, 3H), 7.54-7.53(d, J=4.4 Hz, 1H), 5.21 (s, 2H), 4.06-4.04 (m, 2H), 3.89-3.87 (m, 2H),1.27-1.23 (t, J=6.8 Hz, 3H), 1.11-1.08 (t, J=6.8 Hz, 3H). LC-MS: m/z424.12 (M+H) with a purity of 99%.

Compound 73:2-(8-(dimethylamino)-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Step 1: Preparation of2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(3′-methoxybiphenyl-4-yl)acetamide

Intermediate was prepared using alkylation method A reaction conditionsas described. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 5.19 (s, 2H), 3.74 (s,3H), 3.41 (s, 3H), 3.20 (s, 3H). LC-MS: m/z 332 (M+H).

Step 2: Preparation of methyl2-(8-(dimethylamino)-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetate

A solution of2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(3′-methoxybiphenyl-4-yl)acetamide (1 equiv.) in 11% dimethylamine in 2M ethanolsolution (1 equiv.) was refluxed for 18 h. After consumption of startingmaterial, the reaction mixture was concentrated in vacuo and extractedwith dichloromethane twice. The combined organic layers were washed withbrine, dried over Na₂SO₄ and concentrated under vacuum. The crudecompound was purified by column chromatography to obtain theintermediate methyl2-(8-(dimethylamino)-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetate.TH NMR (400 MHz, DMSO-d₆) δ (ppm): 5.00-4.98 (m, 2H), 3.71 (s, 3H), 3.38(s, 3H), 3.16 (s, 3H), 2.93 (s, 6H). LC-MS: m/z 297 (M+H+41).

Step 3: Preparation of2-(8-(dimethylamino)-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)aceticacid.

Intermediate was prepared using hydrolysis reaction conditions asdescribed. LC-MS: m/z 282-M+H).

Step 4: Preparation of2-(8-(dimethylamino)-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Product prepared using amide coupling method A reaction conditions asdescribed. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.42 (s, 1H), 7.79-7.78(m, 1H), 7.69 (s, 1H), 7.67 (s, 1H), 7.62-7.60 (m, 3H), 7.52 (dd, J=5Hz, J=1 Hz, 1H), 5.02 (s, 2H), 3.41 (s, 3H), 3.17 (s, 3H), 2.97 (s, 6H).LC-MS: m/z 439 (M+H) with a purity of 97%.

Compound 74:2-(1,3-dimethyl-8-morpholino-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Step 1: Preparation of methyl2-(8-bromo-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetate

Intermediate was prepared using alkylation method A reaction conditionsas described. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 5.19 (s, 2H), 3.74 (s,3H), 3.41 (s, 3H), 3.20 (s, 3H). LC-MS: m/z 332 (M+H)

Step 2: Preparation of methyl2-(1,3-dimethyl-8-morpholino-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetate

A stirred solution of methyl2-(8-bromo-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetate(1 equiv.) and morpholine (5 equiv.) in DMF (0.4 M) was refluxed for 2h. After consumption of starting material, the reaction mixture wasquenched with water and extracted with dichloromethane twice. Thecombined organic layers were washed with brine, dried over Na₂SO₄ andconcentrated under vacuum. The crude compound was purified by columnchromatography to afford the intermediate methyl2-(1,3-dimethyl-8-morpholino-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetate.¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 4.94 (s, 2H), 3.71-3.69 (m, 6H), 3.17(s, 3H), 3.39 (s, 3H), 3.16-3.13 (m, 5H). LC-MS: m/z 338 (M+H).

Step 3: Preparation of2-(1,3-dimethyl-8-morpholino-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)aceticacid

Intermediate was prepared using hydrolysis reaction conditions asdescribed. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 4.83 (s, 2H), 3.72-3.70(m, 3H), 3.39 (s, 3H), 3.18 (s, 3H), 3.16-3.14 (m, 5H). LC-MS: m/z 324(M+H).

Step 4: Preparation of2-(1,3-dimethyl-8-morpholino-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Product was prepared using amide coupling method A reaction conditionsas described. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.45 (s, 1H), 7.79 (m,1H), 7.69-7.67 (m, 2H), 7.63-7.60 (m, 3H), 7.52 (m, 1H), 4.97 (s, 2H),3.70 (t, J=4.4 Hz, 4H), 3.42 (s, 3H), 3.21 (t, J=4.4 Hz, 4H), 3.19 (s,3H). LC-MS: m/z 481 (M+H) with a purity of 99%.

Compound 75:2-(1,3-dimethyl-2,6-dioxo-8-(thiophen-2-yl)-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Step 1: Preparation of methyl2-(8-bromo-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetate

Intermediate prepared using alkylation method A reaction conditions asdescribed. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 5.19 (s, 2H), 3.74 (s,3H), 3.41 (s, 3H), 3.20 (s, 3H). LC-MS: m/z 332 (M+H).

Step 2: Preparation of2-(1,3-dimethyl-2,6-dioxo-8-(thiophen-2-yl)-2,3-dihydro-1H-purin-7(6H)-yl)aceticacid

Intermediate prepared using Suzuki coupling method C reaction conditionsas described. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 7.87-7.85 (m, 1H),7.55-7.54 (m, 1H), 7.27-7.24 (m, 1H), 5.28 (s, 2H), 3.47 (s, 3H), 3.24(s, 3H). LC-MS: m/z 321 (M+H).

Step 3: Preparation of2-(1,3-dimethyl-2,6-dioxo-8-(thiophen-2-yl)-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Product prepared using amide coupling method A reaction conditions asdescribed. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.61 (s, 1H), 7.85 (dd,J=5.0 Hz, J=1.2 Hz, 1H), 7.80-7.79 (m, 1H), 7.70-7.68 (m, 2H), 7.62-7.59(m, 4H), 7.54 (dd, J=15.8 Hz, J=1.2 Hz, 1H), 7.25-7.23 (m, 1H), 5.44 (s,2H), 3.49 (s, 3H), 3.24 (s, 3H). LC-MS: m/z 478 (M+H) with a purity of98%.

Compound 76:2-(8-bromo-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Step 1: Methyl2-(8-bromo-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetate

Intermediate prepared using alkylation method A reaction conditions asdescribed. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 5.19 (s, 2H), 3.74 (s,3H), 3.41 (s, 3H), 3.20 (s, 3H). LC-MS: m/z 332 (M+H).

Step 2:2-(8-bromo-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)aceticacid

Intermediate was prepared using hydrolysis reaction conditions asdescribed. ¹H NMR (400 MHz, DMSO-d₆): 13.60 (brs, 1H), 5.06 (s, 2H),3.32 (s, 3H), 3.20 (s, 3H). LC-MS: m/z 316.94, 318.94 M−H, M−H+2) with apurity of 99%.

Step 3:2-(8-bromo-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)acetamide

Product was prepared using amide coupling method A reaction conditionsas described. ¹H NMR (400 MHz, DMSO-d₆): 10.57 (s, 1H), 7.80 (s, 1H),7.70-7.68 (m, 2H), 7.62-7.59 (m, 3H), 7.52 (s, 1H), 5.20 (s, 2H), 3.43(s, 3H), 3.21 (s, 3H). LC-MS: m/z 474.20 (M+H) with a purity of 96.04%.HPLC: At 279 nm with a purity of 97%.

Compound 77:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(6-phenylpyridazin-3-yl)propanamide

Step 1: Preparation of ethyl2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl) propanoate

Intermediate was prepared using Mitsunobu reaction conditions asdescribed. LC-MS: m/z 281 (M+H) with a purity of 46%.

Step 2: Preparation of2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanoic acid

Intermediate was prepared using hydrolysis reaction conditions asdescribed. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 13.27 (br s, 1H), 8.21 (s,1H), 5.49-5.44 (q, J=7.5 Hz, 1H), 3.44 (s, 3H), 3.21 (s, 3H), 1.76-1.74(d, J=7.5 Hz, 3H).

Step 3: Preparation of2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(6-phenylpyridazin-3-yl)propanamide

Product was prepared using amide coupling method A reaction conditionsas described. ¹H-NMR (400 MHz; CDCl₃) δ (ppm): 11.78 (s, 1H), 8.50-8.46(m, 1H), 8.02-8.00 (dd, J=6.8, 3.2 Hz, 2H), 7.90-7.86 (m, 1H), 7.51-7.48(m, 3H), 6.02-5.94 (q, J=7.6 Hz, 1H), 3.61 (s, 3H), 3.44 (s, 3H),1.97-1.95 (d, J=7.2 Hz, 3H). MS (ESI): m/z 406.21 [M+H]+.

Compound 78:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(6-(4-fluorophenyl)pyridazin-3-yl)propanamide

Step 1: Preparation of ethyl2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl) propanoate

Intermediate was prepared using Mitsunobu reaction conditions asdescribed. LC-MS: m/z 281 (M+H) with a purity of 46%.

Step 2: Preparation of2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanoic acid

Intermediate was prepared using hydrolysis reaction conditions asdescribed. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 13.27 (br s, 1H), 8.21 (s,1H), 5.49-5.44 (q, J=7.5 Hz, 1H), 3.44 (s, 3H), 3.21 (s, 3H), 1.76-1.74(d, J=7.5 Hz, 3H).

Step 3: Preparation of2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(6-(4-fluorophenyl)pyridazin-3-yl)propanamide

Product was prepared using amide coupling method A reaction conditionsas described. ¹H-NMR (400 MHz; CDCl₃) δ (ppm): 10.39 (s, 1H), 8.46-8.44(d, J=9.2 Hz, 1H), 8.03-8.00 (m, 2H), 7.88 (s, 1H), 7.84-7.82 (d, J=9.2Hz, 1H), 7.21-7.16 (t, J=8.4 Hz, 2H), 5.91-5.87 (q, J=7.1 Hz, 1H), 3.60(s, 3H), 3.45 (s, 3H), 1.96-1.94 (d, J=7.1 Hz, 3H). MS (ESI): m/z 422[M−H]+.

Compound 79:(S)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)propanamide

Step 1: Preparation of (S)-ethyl2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl) propanoate

Intermediate was prepared using Mitsunobu reaction conditions asdescribed. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 7.76 (s, 1H), 5.62 (q,J=7.6 Hz, 1H), 4.25 (q, J=6.8 Hz, 2H), 3.61 (s, 3H), 3.39 (s, 3H), 1.85(d, J=7.6 Hz, 3H), 1.29 (d, J=7.2 Hz, 3H). MS (ESI): m/z 281 [M+H]+.LC-MS: m/z 281 (M+H) with a purity of 48%.

Step 2: Preparation of(S)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanoicacid

Intermediate was prepared using hydrolysis reaction conditions asdescribed. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 13.28 (brs, 1H), 8.21 (s,1H), 5.47 (q, J=7.6 Hz, 1H), 3.44 (s, 3H), 3.21 (s, 3H), 1.75 (d, J=7.6Hz, 3H). MS (ESI): m/z 253.3 [M+H]+. LC-MS: Purity of 99%

Step 3: Preparation of(S)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(6-phenylpyridazin-3-yl)propanamide

Product was prepared using amide coupling method A reaction conditionsas described. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 11.78 (s, 1H), 8.35 (s,1H), 8.27 (dd, J=9.2 Hz, J=9.2 Hz, 2H), 8.1 (d, J=6.4 Hz, 2H), 7.57-7.51(m, 3H), 5.84 (d, J=7.6 Hz, 1H), 3.46 (s, 3H), 3.19 (s, 3H), 1.9 (d,J=7.6 Hz, 3H). MS (ESI): m/z 406.21 [M+H]+. LC-MS: Purity of 97%.

Compound 80:(R)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiophen-3-yl)phenyl)propanamide

Step 1: Preparation of (R)-ethyl2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7 (6H)-yl) propanoate

Intermediate was prepared using Mitsunobu reaction conditions asdescribed. ¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 7.76 (s, 1H), 5.62 (q,J=7.6 Hz, 1H), 4.25 (q, J=6.8 Hz, 2H), 3.61 (s, 3H), 3.39 (s, 3H), 1.85(d, J=7.6 Hz, 3H), 1.29 (d, J=7.2 Hz, 3H). MS (ESI): m/z 281 [M+H]+.LC-MS: m/z 281 (M+H) with a purity of 48%.

Step 2: Preparation of(R)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl) propanoicacid

Intermediate prepared using hydrolysis reaction conditions as described.¹H-NMR (400 MHz; DMSO-d₆) δ (ppm): 13.28 (brs, 1H), 8.21 (s, 1H), 5.47(q, J=7.6 Hz, 1H), 3.44 (s, 3H), 3.21 (s, 3H), 1.75 (d, J=7.6 Hz, 3H).MS (ESI): m/z 253.3 [M+H]+. LC-MS: Purity of 99%

Step 3: Preparation of(R)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(6-phenylpyridazin-3-yl)propanamide

Product was prepared using amide coupling method A reaction conditionsas described. ¹H-NMR (400 MHz; CDCl₃) δ (ppm): 10.61 (s, 1H), 8.47 (d,J=9.6 Hz, 1H), 8.01 (dd, J=2.4 Hz, J=5.6 Hz, 2H), 7.89 (s, 1H), 7.86 (s,1H), 7.49 (d, J=6.8 Hz, 3H), 5.98 (d, J=6.8 Hz, 1H), 3.61 (s, 3H), 3.43.(s, 3H), 1.96 (d, J=7.6 Hz, 3H). MS (ESI): m/z 406.21 [M+H]+. LC-MS:Purity of 97%.

Compound 81:2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiazol-2-yl)phenyl propanamide

Step 1: Preparation of ethyl2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl) propanoate

Intermediate was prepared using alkylation method A reaction conditionsas described. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 8.20 (s, 1H), 5.55 (q,J=7.2 Hz, 1H), 4.14 (q, J=7.2 Hz, 2H), 3.45 (s, 3H), 3.21 (s, 3H), 1.75(d, J=7.2 Hz, 3H), 1.17 (t, J=7.2 Hz, 3H). LC-MS: m/z 281 (M+H)

Step 2: Preparation of 2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanoic acid

Intermediate was prepared using hydrolysis reaction conditions asdescribed. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 8.20 (s, 1H), 5.47 (q,J=7.2 Hz, 1H), 3.44 (s, 3H), 3.21 (s, 3H), 1.75 (d, J=7.2 Hz, 3H).LC-MS: m/z 253 (M+H)

Step 3: Preparation of2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4-(thiazol-2-yl)phenyl)propanamide

Product was prepared using amide coupling method D reaction conditionsas described. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 10.65 (s, 1H), 8.33 (s,1H), 7.93-7.91 (m, 2H), 7.88 (d, J=3.2 Hz., 1H), 7.73-7.72 (m, 2H), 7.70(m, 1H), 5.73-5.68 (q, J=7.2 Hz, 1H), 3.46 (s, 3H), 3.20 (s, 3H), 1.85(d, J=7.2 Hz, 3H). LC-MS: m/z 411 (M+H) with a purity of 99%.

Compound 82: 2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(6-(thiazol-2-yl) pyridin-3-yl)propanamide

Step 1: Preparation of ethyl2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl) propanoate

Intermediate was prepared using alkylation method A reaction conditionsas described. ¹H NMR (400 MHz, CDCl₃) δ (ppm): 7.97 (s, 1H), 5.59 (q,J=7.60 Hz, 1H), 4.24 (q, J=7.20 Hz, 2H), 3.58 (s, 3H), 3.36 (s, 3H),1.83 (d, J=7.60 Hz, 3H), 1.28 (t, J=7.20 Hz, 3H). LC-MS: m/z 281 (M+H).

Step 2: Preparation of2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)propanoic acid

Intermediate was prepared using hydrolysis reaction conditions asdescribed. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 8.19 (s, 1H), 5.46 (q,J=7.20 Hz, 1H), 3.44 (s, 3H), 3.21 (s, 3H), 1.74 (d, J=7.20 Hz, 3H).LC-MS: m/z 253 (M+H).

Step 3: Preparation of2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(6-(thiazol-2-yl)pyridin-3-yl)propanamide

Product was prepared using amide coupling method E reaction conditionsas described. ¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 11.30 (s, 1H), 8.93 (s,1H), 8.30-8.32 (m, 2H), 8.09-8.11 (m, 1H), 7.95 (d, J=2.00 Hz, 1H), 7.82(d, J=2.00 Hz, 1H), 5.80 (q, J=7.20 Hz, 3H), 3.46 (s, 3H), 3.19 (s, 3H),1.86 (d, J=7.20 Hz, 3H). LC-MS: m/z 412 (M+H) with a purity of 97%.

Compound 83:(R)-2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(5-(thiazol-2-yl)pyridin-2-yl)propanamide

Product was prepared using chiral separation of the racemate. ¹H NMR(400 MHz, DMSO-d₆) 3 (ppm): 11.30 (s, 1H), 8.93 (s, 1H), 8.30-8.32 (m,2H), 8.09-8.11 (m, 1H), 7.95 (d, J=2.00 Hz, 1H), 7.82 (d, J=2.00 Hz,1H), 5.80 (q, J=7.20 Hz, 3H), 3.46 (s, 3H), 3.19 (s, 3H), 1.86 (d,J=7.20 Hz, 3H).

Compound 84:(S)-2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(5-(thiazol-2-yl)pyridin-2-yl)propanamide

Product was prepared using chiral separation of the racemate. ¹H NMR(400 MHz, DMSO-d₆) δ (ppm): 11.30 (s, 1H), 8.93 (s, 1H), 8.30-8.32 (m,2H), 8.09-8.11 (m, 1H), 7.95 (d, J=2.00 Hz, 1H), 7.82 (d, J=2.00 Hz,1H), 5.80 (q, J=7.20 Hz, 3H), 3.46 (s, 3H), 3.19 (s, 3H), 1.86 (d,J=7.20 Hz, 3H).

Compound 85:(R)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(2-methylthiazol-4-yl)pyridin-2-yl)propanamide

Product was prepared using amide coupling method A reaction conditionsas described. The isomers were separated by chiral preparative HPLC togive(R)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(2-methylthiazol-4-yl)pyridin-2-yl)propanamide,¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 11.18 (1H, s), 8.94-8.93 (1H, d,J=2.4 Hz), 8.33 (1H, s), 8.30-8.27 (1H, dd, J1=2.4 Hz, J2=6.4 Hz),8.05-8.03 (1H, d, J=8.8 Hz), 8.01 (1H, s), 5.80-5.78 (1H, m) 3.45 (3H,s), 3.19 (3H, s), 2.72 (3H, s), 1.86-1.84 (3H, d, J=7.6 Hz). LC-MS: m/z426.12 (M+H) with a purity of 97.84%. HPLC: At 254 nm with a purity of97%. Chiral HPLC: 99%. Specific Rotation: +137 deg

Compound 86:(S)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(2-methylthiazol-4-yl)pyridin-2-yl)propanamide

Product was prepared using amide coupling method A reaction conditionsas described. The isomers were separated by chiral preparative HPLC togive 100 mg of(S)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5-(2-methylthiazol-4-yl)pyridin-2-yl)propanamide.

¹H NMR (400 MHz, DMSO-d₆) δ (ppm): 11.18 (s, 1H), 8.94-8.93 (d, J=2.4Hz, 1H), 8.33 (s, 1H), 8.30-8.27 (dd, J1=2.4 Hz, J2=6.4 Hz, 1H),8.05-8.03 (d, J=8.8 Hz, 1H), 8.01 (s, 1H), 5.80-5.78 (q, J=7.5 Hz, 1H)3.45 (s, 3H), 3.19 (s, 3H), 2.72 (s, 3H), 1.86-1.84 (d, J=7.6 Hz, 3H).LC-MS: m/z 426.12 (M+H) with a purity of 98.38%. HPLC: At 254 nm with apurity of 97%. Chiral HPLC: 99%. Specific Rotation: −127 deg

Compound 87:3-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(4-(thiophen-3-yl)phenyl)propanamide

Step 1: Preparation of ethyl3-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)propanoate

The intermediate was prepared using alkylation conditions. ¹H NMR (400MHz, DMSO-d₆): 8.03 (s, 1H), 4.46 (t, J=6.8 Hz, 2H), 4.05 (q, t=6.6 Hz,2H), 3.42 (s, 3H), 3.24 (s, 3H), 2.93 (t, J=6.6 Hz, 2H), 1.15 (t, J=6.8Hz, 3H). LC-MS: m/z 281 (M+H).

Step 2: Preparation of3-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)propanoicacid

¹H NMR (400 MHz, DMSO-d₆): 8.01 (s, 1H), 4.43 (t, J=6.8 Hz, 2H), 3.42(s, 3H), 3.24 (s, 3H), 2.84 (t, J=6.8 Hz, 2H. LC-MS: m/z 253 (M+H)

Step 3: Preparation of3-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(4-(thiophen-3-yl)phenyl)propanamide

¹H NMR (400 MHz, DMSO-d₆): 9.99 (s, 1H), 8.00 (s, 1H), 7.75 (br s, 1H),7.64-7.55 (m, 5H), 7.51-7.49 (d, J=5.2 Hz, 1H), 4.54 (t, J=6.4 Hz, 2H),3.42 (s, 3H), 2.96 (t, J=6.4 Hz, 2H). LC-MS: m/z 410 (M+H) with a purityof 99%.

Compound 88:4-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(4-(thiophen-3-yl)phenyl)butanamide

Step 1: Preparation of ethyl4-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)butanoate

The intermediate was prepared using alkylation conditions. ¹H NMR (400MHz, CDCl₃-d) δ 7.55 (s, 1H), 4.37 (t, J=6.80 Hz, 2H), 4.13 (q, J=7.20Hz, 2H), 3.59 (s, 3H), 3.41 (s, 3H), 2.32 (t, J=6.80 Hz, 2H), 2.21 (p,J=6.80 Hz, 2H), 1.25 (q, J=7.20 Hz, 3H). LC-MS: m/z 295 (M+H) with apurity of 98%.

Step 2: Preparation of4-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)butanoic acid

The intermediate was prepared using hydrolysis conditions. ¹H NMR (400MHz, DMSO-d₆) δ 8.05 (s, 1H), 4.26 (t, J=7.20 Hz, 2H), 3.42 (s, 3H),3.22 (s, 3H), 2.19 (t, J=7.20 Hz, 2H), 2.01 (q, J=7.20 Hz, 2H). LC-MS:m/z 267 (M+H).

Step 3: Preparation of4-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(4-(thiophen-3-yl)phenyl)butanamide

The final product is prepared using amide coupling method A reactionconditions. ¹H NMR (Methanol-d₄) δ 7.92 (s, 1H), 7.53-7.57 (m, 3H),7.47-7.50 (m, 2H), 7.43-7.45 (m, 1H), 7.40-7.41 (m, 1H), 4.45 (t, J=6.80Hz, 2H), 3.45 (s, 3H), 3.33 (s, 3H), 2.43 (t, J=6.80 Hz, 2H), 2.31 (p,J=6.80 Hz, 2H). LCMS (ESI) m/z 424 (MH+) with a purity of 97%.

Compound 89:2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(4-(thiophen-2-yl)phenyl)acetamide

¹H NMR (400 MHz, DMSO-d₆): 10.48 (s, 1H), 8.07 (s, 1H), 7.62 (s, 4H),7.49 (d, J=5.2 Hz, 1H), 7.43 (d, J=3.6 Hz, 1H), 7.11 (m, 1H), 5.22 (s,2H), 3.47 (s, 3H), 3.21 (s, 3H). LC-MS: m/z 396 (M+H) with a purity of97%.

Compound 90:3-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(4-(thiophen-2-yl)phenyl)propanamide

¹H NMR (400 MHz, DMSO-d₆): 10.05 (s, 1H), 8.00 (s, 1H), 7.57 (s, 4H),7.47 (d, J=5.2 Hz, 1H), 7.40 (d, J=3.6 Hz, 1H), 7.10 (t, J=4.2 Hz, 1H),4.54 (t, J=6.4 Hz, 2H), 3.42 (s, 3H), 3.26 (s, 3H), 2.96 (t, J=6.4 Hz,2H). LC-MS: m/z 410 (M+H) with a purity of 97%.

Compound 91:4-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(4-(thiophen-2-yl)phenyl)butanamide

¹H NMR (Methanol-d₄) δ 7.92 (s, 1H), 7.47-7.55 (m, 4H), 7.30-7.32 (m,2H), 7.06 (t, J=4.40 Hz, 1H), 4.45 (t, J=6.80 Hz, 2H), 3.45 (s, 3H),3.33 (s, 3H), 2.43 (t, J=6.80 Hz, 2H), 2.30 (p, J=6.80 Hz, 2H). LCMS(ESI) m/z 424 (MH+). LCMS (ESI) m/z 424 (MH+) with a purity of 98%.

Compound 92:3-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(4-(2-methylthiazol-4-yl)phenyl)propanamide

1H NMR (400 MHz, Methanol-d₄) δ 7.92 (s, 1H), 7.78-7.80 (m, 2H),7.54-7.55 (m, 3H), 4.67 (t, J=6.00 Hz, 2H), 4.55 (bs, 1H), 3.52 (s, 3H),3.37 (s, 3H), 3.01 (t, J=6.00 Hz, 2H), 2.73 (s, 3H). LC-MS: m/z 425(M+H) with a purity of 99%.

Compound 93:3-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(4-(thiazol-2-yl)phenyl)propanamide

¹H NMR (400 MHz, DMSO-d₆) δ 10.17 (s, 1H), 8.00 (s, 1H), 7.85-7.88 (m,3H), 7.65-7.70 (m, 3H), 4.54 (t, J=6.40 Hz, 2H), 3.42 (s, 3H), 2.99 (t,J=6.40 Hz, 2H). LC-MS: m/z 411 (M+H) with a purity of 99%.

Compound 94:4-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(4-(thiazol-2-yl)phenyl)butanamide

¹H NMR (400 MHz, Methanol-d₄) δ 7.95 (s, 1H), 7.86 (d, J=8.80 Hz, 2H),7.82 (d, J=3.20 Hz, 1H), 7.61 (d, J=8.80 Hz, 2H), 7.55 (d, J=3.20 Hz,1H), 4.45 (t, J=6.80 Hz, 2H), 3.45 (s, 3H), 3.33 (s, 3H), 2.45 (t,J=6.80 Hz, 2H), 2.31 (p, J=6.80 Hz, 2H). LC-MS: m/z 425 (M+H) with apurity of 96%.

Application Example 1

Materials and Methods:

Cell Lines and Culture Conditions:

HEK293-STF cell line was modified from Human embryonic kidney cell lineHEK293 transfected with the STF reporter. HEK293-STF3A cell line wasfurther modified from HEK293-STF cell line to express Wnt3A. This cellline was used to identify compounds that regulate either early or latesignalling components of the Wnt pathway. L-Wnt3A (ATCC, #CRL-2647) cellline was used for providing Wnt3A conditioned media. The three celllines were grown in DMEM (Dulbecco's Modified Eagle Medium) with 10% FBS(fetal bovine serum) incubated in 37° C. with 5% CO₂.

Cell Viability Assay:

5000 cells in 75 μl culture media were seeded in each well of black 96well plates (Greiner #655090) and incubated overnight at 37° C. 25 μl ofserially diluted compound was added to the cells giving finalconcentration of 50 μM to 1.5 nM. After 1 day of treatment, 100 μl ofCellTiter-Glo® Luminescent Cell Viability Assay reagent (#G7571,Promega) was added to each well and incubated for 10 minutes at roomtemperature. Luminescence was measured using Tecan Safire2® microplatereader.

STF3A Assay:

2×10⁴ HEK293-STF3A cells in 75 μl culture media were seeded in each wellof white 96 well plates (Greiner #655098) and incubated overnight at 37°C. 25 μl serially diluted compound was added to the cells to give finalconcentration of 50 μM to 1.5 nM. After 1 day of treatment, 100 μl ofSteady-Glo® Luciferase Assay reagent (#E2520, Promega) was added to eachwell and incubated for 10 minutes at room temperature. Luminescence wasmeasured using Tecan Safire2® plate reader.

STF/WNT3A Conditioned Medium (STF/WNT3A CM) Assay:

L-Wnt3A cells were cultured in three T-175 flasks at 3×10⁴ cells/ml in30 ml culture medium per flask. After 4 days of incubation, the Wnt3Aconditioned media were harvested and then centrifuged at 2000 rpm for 10minutes to remove the debris. The Wnt3A conditioned media were stored at−20° C. if not used immediately.

2×104 HEK293-STF cells in 25 μl culture media were added in each well ofwhite 96 well plates (Greiner #655098). 25 μl serially diluted compoundwas added to the cells. After 4 hours of incubation, 100 μl Wnt-3Aconditioned medium was added to the cells. The final concentration ofcompound ranged from 33 μM to 1 nM. After incubation for 1 day at 37°C., 100 μl of Steady-Glo® Luciferase Assay reagent (#E2520, Promega) wasadded to each well and incubated for 10 minutes at room temperature.Luminescence was measured using Tecan Safire2® microplate reader.

Results:

Compound STF3A IC50 μM 1 <0.1 2 <0.1 3 <1 4 <1 5 <0.1 6 <1 7 >10 8 <5 9<0.1 10 <1 11 <0.1 12 <5 13 >10 14 <1 15 <0.1 16 <1 17 >10 18 <0.1 19<0.1 20 <0.1 21 <0.1 22 <0.1 23 <0.1 24 <1 25 <0.1 26 <0.1 27 >10 28 >1029 <0.1 30 <0.1 31 <5 32 <0.1 33 <0.1 34 <0.1 35 <0.1 36 <0.1 37 <0.1 38<0.1 39 >10 40 <1 41 <0.1 42 >10 43 >10 44 >10 45 <0.1 46 <1 47 <1 48 <149 <1 50 <1 51 <1 52 <1 53 <0.1 54 <0.1 55 <5 56 <1 57 <0.1 58 >10 59<0.1 60 <1 61 <0.1 62 <0.1 63 <1 64 <0.1 65 >10 66 <0.1 67 <1 68 <0.1 69<0.1 70 <0.1 71 >10 72 <0.1 73 <0.1 74 <1 75 <1 76 <0.1 77 <0.1 78 <1 79<0.1 80 >10 81 <0.1 82 <0.1 83 <10 84 <0.1 85 <5 86 <0.1 87 <0.1 88 <0.189 <0.1 90 <0.1 91 <0.1 92 <1 93 <1 94 <1

MMTV-WNT1 Tumor Model:

To test the in vivo efficacy of Compound 5 to prevent the growth of Wntdriven tumors, fragments from two independent MMTV-WNT1 tumors wereorthotopically transplanted into female nude mice. The mice were treatedwith either vehicle or Compound 5, 30 mg/kg once daily for 19 days.Tumor volumes were measured on alternate days. Treatment with Compound 5decreased tumor growth in all the treated mice. A significant decreasein tumor weights collected at sacrifice was also observed. Results areshown in FIG. 1.

Cytoplasmic and Nuclear β-Catenin Experiment

Result: Compound 5 decreased cytoplasmic and nuclear β-catenin intumors. Staining the tumor sections for β-catenin showed that vehicletreated tumors had abundant β-catenin in cytoplasm and nucleus. Tworepresentative samples from each treatment arm are shown in FIG. 2.

Phospho-LRP6 Assay as a Target Efficacy Marker for Compound 5

Palmitoylation of Wnts is essential for Wnt/β-Catenin signaling. OnceWnts are palmitoylated by the O-acyl transferase porcupine they aresecreted and subsequently bind to the receptor complex, consisting ofFrizzled (cognate receptors) and the co-receptor LRP5 or LRP6 (Cadiganand Peifer, 2009). LRP5 and LRP6 are highly homologous single-passtransmembrane proteins of the low-density lipoprotein receptor(LDLR)-related protein family. Upon Wnt binding LRP is phosphorylated onmultiple sites (including Thr 1479, Ser 1490 and Thr1493) by kinasessuch as Casein Kinase 1 (CK1), Glycogen Synthase Kinase 3 (GSK3) or MEK1(Cervenka et al., 2011; Tamai et al., 2004; Zeng et al., 2005).Phosphorylated LRP then recruits axin to the membrane and subsequentlyactivates β-Catenin signalling.

The present target efficacy biomarker assay measures a decrease inlevels of p-LRP6 (i.e. phosphorylated LRP-6) (Ser1490) upon treatmentwith a porcupine inhibitor (Compound 5). Cells treated with 2 μM of thetest compound in vitro showed a greater than 50% reduction in p-LRP6from 4 h onwards, with no decrease observed in total LRP levels (FIG.3). The inhibitory effect remains up to 72 h (data not shown) in thepresence of the compound and up to 12 h after the compound has beenremoved (data not shown).

While 2 μM led to 50-60% inhibition after 6 h of in vitro treatment, 3.3nM of Compound 5 still inhibited p-LRP6 (Ser1490) by about 20% in thisassay (FIG. 4). Concentrations lower than 3.3 nM did not inhibit p-LRP6in HPAF-II pancreatic adenocarcinoma cells (data not shown).

The inventors have validated that this assay works alike for cancercells and tumour tissue (data not shown).

REFERENCES

-   Cadigan, K. M., and Peifer, M. (2009). Wnt signaling from    development to disease: insights from model systems. Cold Spring    Harbor perspectives in biology 1, a002881.-   Cervenka, I., Wolf, J., Masek, J., Krejci, P., Wilcox, W. R.,    Kozubik, A., Schulte, G., Gutkind, J. S., and Bryja, V. (2011).    Mitogen-activated protein kinases promote WNT/beta-catenin signaling    via phosphorylation of LRP6. Molecular and cellular biology 31,    179-189.-   Tamai, K., Zeng, X., Liu, C., Zhang, X., Harada, Y., Chang, Z., and    He, X. (2004). A mechanism for Wnt coreceptor activation. Molecular    cell 13, 149-156.-   Zeng, X., Tamai, K., Doble, B., Li, S., Huang, H., Habas, R.,    Okamura, H., Woodgett, J., and He, X. (2005). A dual-kinase    mechanism for Wnt co-receptor phosphorylation and activation. Nature    438, 873-877.

1-66. (canceled)
 67. A compound having structure (I)

wherein: R¹, R², R³, R⁴ and R⁵ are each, independently, H or an alkylgroup; D is selected from the group consisting of H, halogen, alkyl,cycloalkyl, aryl, and dialkylamino, each (other than H and halogen)being optionally substituted; Ar is a 6-membered heteroaryl ring having1 or 2 nitrogen atoms having no substituents other than the Cy moietyand amide nitrogen atom depicted in structure (I), these being in a1,4-relationship on the ring; and Cy is an aryl, heteroaryl, or asaturated ring containing at least one heteroatom, each being optionallysubstituted; and n is an integer from 1 to 2; or an enantiomer,diastereomer, or salt thereof.
 68. The compound of claim 67, wherein R¹and R² are either both methyl or both ethyl.
 69. The compound of claim67, wherein n is
 1. 70. The compound of claim 67, wherein D is selectedfrom H, methyl, cyclopropyl, trifluoromethyl, phenyl, dimethylamino,morpholin-N-yl, thiophene-3-yl, and bromo.
 71. The compound of claim 67,wherein D is H.
 72. The compound of claim 67, wherein R⁵ is H.
 73. Thecompound of claim 67, wherein one of R³ and R⁴ is H and the other isselected from H, methyl, and ethyl.
 74. The compound of claim 67,wherein Ar is selected from the following:

wherein

denotes the point of connection to the amide nitrogen atom depicted instructure (I) and * denotes the point of connection to the Cy moietydepicted in structure (I).
 75. The compound of claim 67, wherein Cy is a5- or 6-membered aromatic ring having between 0 and 2 nitrogen atoms, 0or 1 sulfur atoms, and 0 or 1 oxygen atoms.
 76. The compound of claim67, wherein Cy is selected from the following:

wherein * denotes the point of connection to the Ar moiety of structure(I).
 77. The compound of claim 67, selected from:


78. An anhydrous form of2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(6-phenylpyridazin-3-yl)acetamidefree base, as represented by:


79. The anhydrous form of claim 78, which is a non-hydrated singlepolymorph.
 80. The anhydrous form of claim 78, which is crystalline. 81.The anhydrous form of claim 80, which shows on X-ray diffraction a peakon the 2theta scale at 22.2°±0.5°.
 82. The anhydrous form of claim 80,which shows on X-ray diffraction peaks on the 2theta scale at 5.5°±0.5°and 14.2°±0.5°.
 83. The anhydrous form of claim 80, which shows on X-raydiffraction at least four peaks on the 2theta scale selected from5.5°±0.5°, 12.5°±0.5°, 14.2°±0.5°, 16.7°±0.5°, 17.7°±0.5°, 18.8°±0.5°,22.4°±0.5°, 24.2°±0.5° and 31.7°±0.5°.
 84. A pharmaceutical compositioncomprising a compound according to claim 67 together with one or morepharmaceutically acceptable carriers, diluents or adjuvants.
 85. Amethod of treating a disease or condition associated with Wnt pathwayactivity, comprising administering to a subject in need thereof atherapeutically effective amount of a compound according to claim 67,wherein the disease or condition is selected from the cancer, pulmonaryfibrosis, liver fibrosis, skin fibrosis, renal fibrosis, a degenerativedisease, diabetic retinopathy, stem cell retinopathy, rheumatoidarthritis, psoriasis, and myocardial infarction.
 86. The method of claim85, wherein the disease or condition is a cancer selected from cervical,colon, breast, bladder, head and neck, gastric, lung, ovarian, prostate,thyroid, non-small-cell lung, chronic lymphocytic leukemia,mesothelioma, melanoma, pancreatic adenocarcinoma, basal cell carcinoma,osteosarcoma, hepatocellular carcinoma, Wilm's tumor, andmedulloblastoma.